Amines for detection of dopamine by generation of hydrogen peroxide and peroxyoxalate chemiluminescence

A range of nitrogen-containing compounds (alkyl amines, piperazines, cyclohexylamines and nitrogen heterocyclics) were investigated for generation of hydrogen peroxide from dopamine and detection by peroxyoxalate chemiluminescence. Imidazole, ethyleneurea and allantoin among the nitrogen heterocyclic compounds tested generated hydrogen peroxide from dopamine following incubation at 60°C, pH 9.5–10.5, for 0–30 min. Imidazole was the most effective for generation of hydrogen peroxide, but imidazole derivatives with a primary amine side chain (histamine) or thiol (ethylenethiourea) were not effective. The presence of a ketone group (ethyleneurea, allantoin) did not hinder the reaction. Under optimal conditions (30 min incubation, 50 mmol/L imidazole) 10.5 nmol of dopamine could be detected. The cyclohexylamines tested produced low amounts of hydrogen peroxide (0.09–2.74% of light intensity with imidazole), and the piperazines and the alkyl amines tested produced no detectable hydrogen peroxide. Imidazole reacts with the phenolic groups of dopamine in a different manner from monoamine oxidase, and a reagent containing imidazole, ethyleneurea or allantoin was useful for non-enzymatic detection of dopamine by peroxyoxalate chemiluminescence.© John Wiley & Sons, Ltd.     

A two-intermediate model for imidazole-promoted peroxyoxalate chemiluminescence

A mechanism is proposed for imidazole-catalysed peroxyoxalate chemiluminescence. The reaction model includes a sequential formation of 1-aroxalylimidazole and 1,1′-oxalyldiimidazole as light-producing reaction intermediates. The suggestion is supported by the kinetic data obtained for the reaction of imidazole with bis(4-nitrophenyl) oxalate and on the recently reported ability of 1,1′-oxalyldiimidazole to function as an efficient chemiluminescence reagent. The relative contributions of different catalytic pathways and hydrolytic side-reactions are discussed © 1997 John Wiley & Sons, Ltd.     

Unprecedented chemiluminescence behaviour during peroxyoxalate chemiluminescence of oxalates with fluorescent or electron-donating aryloxy groups.

A series of diaryl and bis(4-styrylphenyl) oxalates with electron-donating substituents or fluorescent moieties were subjected to the peroxyoxalate chemiluminescence (PO-CL) reaction, some of which were found to behave in a unprecedented manner. The reaction of bis(p-methyoxyphenyl) oxalate, as a representative example, emits light due not only to the emission from the externally added excited fluorophore, but also from the presumable excimer of p-methoxyphenol. Also, during the reaction of the bis(4-styrylphenyl) oxalates, the emission based on the fluorescence as well as the excimer of the eliminating group were observed. These experimental results suggest that such emitting species would be formed by an intra- and intermolecular electronic interaction with a high-energy intermediate, such as a dioxetanone.     

A lab‐on‐a‐chip device for analysis of amlodipine in biological fluids using peroxyoxalate chemiluminescence system

A highly sensitive, rapid and economical method for the determination of amlodipine (AM) in biological fluids was developed using a peroxyoxalate chemiluminescence (CL) system in a lab‐on‐a‐chip device. Peroxyoxalate‐CL is an indirect type of CL that allows the detection of native fluorophores or compounds derivatized with fluorescent labels. Here, fluorescamine was reacted with AM, and the derivatization product was used in a bis‐(2,4,6‐trichlorophenyl)oxalate‐CL system. Fluorescamine reacts selectively with aliphatic primary amine at neutral or basic pH. As most of the calcium channel blocker and many cardiovascular drugs do not contain primary amine, the developed method is highly selective. The parameters that influenced the CL signal intensity were studied carefully. These included the chip geometry, pH, concentration of reagents used and flow rates. Moreover, we confirmed our previous observation about the effects of imidazole, which is commonly used in the bis‐(2,4,6‐trichlorophenyl)oxalate‐CL system as a catalyst, and found that the signal was significantly improved when imidazole was absent. Under optimized conditions, a calibration curve was obtained with a linear range (10–100 µg/L). The limit of detection was 3 µg/L, while the limit of quantification was 10 µg/L. Finally the method was applied for the determination of AM in biological fluids successfully. Copyright © 2014 John Wiley & Sons, Ltd.     

Kinetic studies on the peroxyoxalate chemiluminescence reaction: determination of the cyclization rate constant

Although more currently utilized as analytical tool because of its high sensitivity and good reproducibility, the mechanism of the peroxyoxalate system, a chemiluminescence reaction with quantum yields only comparable to bioluminescence systems, has been extensively studied. The light emission mechanism can be divided in the pathway before chemiexcitation, which contains the rate‐limiting steps, and the fast and kinetically non‐observable chemiexcitation step. In this work, we obtain information on the mechanism of the slow pathways, attribute values to several rate constants prior to chemiexcitation and suggest a mechanistic scheme that could help optimization of conditions when the peroxyoxalate reaction is used as analytical tool. Copyright © 2002 John Wiley & Sons, Ltd.     

Photographic detection of fluorescent-labelled oligodeoxynucleotide in the blotting format by peroxyoxalate chemiluminescence

The preparation of a fluorescent labelled oligonucleotide and its photographic detection by peroxyoxalate chemiluminescence (PO-CL) are described. Fluorescent labelling of an oligonucleotide (15-mer) was performed with naphthalene-2,3-dicarboxaldehyde to give an N-substituted 1-cyanobenz[f]isoindole (CBI) derivative (CBI-15-mer). For the photographic detection of CBI-15-mer, the bis(2,6-difluorophenyl) oxalate (DFPO)-dimethyl phthalate (DMP) system was selected to obtain a long-lived CL emission. After optimizing the conditions for the CL reaction, the system was applied to the photographic detection, and as little as 250 fmol per spot of CBI-15-mer on a membrane were detected as a visible spot with an instant photographic film. © 1998 John Wiley & Sons, Ltd.     

Light from maillard reaction: Photon counting,emission spectrum,photography and visual perception

Several authors have reported on high-sensitivity measurement of oxygen-dependent low-level chemiluminescence (CL) from Maillard reactions (MR), i.e. nonenzymatic amino-carbonyl reactions between reducing sugars and amino acids (also referred to as nonenzymatic browning). Here we report for the first time, that light from Maillard reactions can be seen by the human eye and also can be photographed. In parallel with visual perception and photography CL was monitored by means of a CL-detection programme of a liquid scintillation counter (LSC, single photon rate counting). CL emission spectrum was recorded by a monochromator-microchannel plate photomultiplier arrangement. CL intensity from reaction of 6-aminocaproic acid with D-ribose (200 mg each) in 5 mL H₂O at pH 11 at 95°C was high enough for visual perception after adaptation to absolute darkness. Reaction in dimethylsulphoxide (DMSO) exhibited strongly enhanced CL (10 mg each in 5 mL were sufficient for visual detection) and could be photographed (15 minutes' exposure, ASA 6400); all characteristics of Maillard specific CL (O₂-dependence, no CL from nonreducing sugars, inhibition by sulphur compounds) remained. Visual detection of CL and measurement by LSC were in full concordance. The CL emission spectrum showed two broad peaks at around 500 nm and 695 nm. Fluorescence emission of the brown reaction mixture matched the bluegreen part of the CL emission spectrum. Emission of visible light during Maillard reactions may partly originate from oxygen-dependent generation of excited states and energy transfer to simultaneously formed fluorescent products of the browning reaction.     

Comparison of the ability of ferric complexes to catalyze microsomal chemiluminescence, lipid peroxidation, and hydroxyl radical generation

The interaction of microsomes with iron and NADPH to generate active oxygen radicals was determined by assaying for low level chemiluminescence. The ability of several ferric complexes to catalyze light emission was compared to their effect on microsomal lipid peroxidation or hydroxyl radical generation. In the absence of added iron, microsomal light emission was very low; chemiluminescence could be enhanced by several cycles of freeze-thawing of the microsomes. The addition of ferric ammonium sulfate, ferric-citrate, or ferric-ADP produced an increase in chemiluminescence, whereas ferric-EDTA or -diethylenetriaminepentaacetic acid (detapac) were inhibitory. The same response to these ferric complexes was found when assaying for malondialdehyde as an index of microsomal lipid peroxidation. In contrast, hydroxyl radical generation, assessed as oxidation of chemical scavengers, was significantly enhanced in the presence of ferric-EDTA and -detapac and only weakly elevated by the other ferric complexes. Ferric-desferrioxamine was essentially inert in catalyzing any of these reactions. Chemiluminescence and lipid peroxidation were not affected by superoxide dismutase, catalase, or competitive hydroxyl radical scavengers whereas hydroxyl radical production was decreased by the latter two but not by superoxide dismutase. Chemiluminescence was decreased by the antioxidants propylgallate or glutathione and by inhibiting NADPH-cytochrome P-450 reductase with copper, but was not inhibited by metyrapone or carbon monoxide. The similar pattern exhibited by ferric complexes on microsomal light emission and lipid peroxidation, and the same response of both processes to radical scavenging agents, suggests a close association between chemiluminescence and lipid peroxidation, whereas both processes can be readily dissociated from free hydroxyl radical generation by microsomes.     

Effect of different inhibitors on the intracellularly and extracellularly generated chemiluminescence induced by formylmethionyl-leucyl-phenylalanine in polymorphonuclear leukocytes. Cellular response in the presence of mannitol,benzoate, taurine,indomethacin and NDGA

When polymorphonuclear leukocytes (PMNL) interact with the soluble stimulus formylmethionyl-leucyl-phenylalanine (FMLP), the cells increase their production of oxidative metabolites. This increased production can be measured as lumino-amplified light emission or chemiluminescence (CL). In the present report, experimental systems which allow a quantitation of extracellularly and intracellularly generated metabolites have been used, and the effect of mannitol, benzoate, taurine, indomethacin and nordihydroguaiaretic acid has been investigated. The presence of the hypochlorous acid scavenger taurine had no effect on the intracellular response, whereas the extracellular response was reduced with around 50%. The hydroxyl radical scavenger mannitol had only minor effects on the response, whereas benzoate, another hydroxyl radical scavenger, reduced the extracellular response with around 50% and the intracellular response with more than 90%. Indomethacin, an inhibitor of arachidonic acid metabolism, did not influence the response, whereas NDGA, also an inhibitor of the arachidonic acid metabolism, totally abolished both the extracellular and the intracellular response. The use of scavengers/inhibitors as a means of determining the mechanisms of light emission, and the origin of chemiluminescence produced by neutrophils stimulated by FMLP is discussed.     

Chemiluminescence studies on the generation of oxygen radicals from the interaction of NADPH-cytochrome P-450 reductase with iron

The ability of NADPH-cytochrome P-450 reductase to interact with iron and generate oxygen radicals was evaluated by assaying for low level chemiluminescence. The basic reaction system which contained the reductase, an NADPH-generating system, ferric-EDTA as an electron acceptor, and t-butyl hydroperoxide as the oxidant acceptor, resulted in the production of chemiluminescence. Omission of any of these components resulted in a complete loss of chemiluminescence. The light emission was completely sensitive to inhibition by glutathione and butylated hydroxytoluene, partially sensitive (about 60% decrease) to catalase and hydroxyl radical scavengers, and relatively insensitive (about 20% decrease) to superoxide dismutase. The ability of other ferric chelates to replace ferric-EDTA in catalyzing the reductase-dependent chemiluminescence was evaluated. Ferric-citrate, -ADP, -ATP, and ferric-ammonium sulfate were ineffective in promoting chemiluminescence, whereas ferric-diethylenetriaminepentaacetic acid was even more effective than ferric-EDTA. Thus, the ferric chelates, which catalyze reductase-dependent chemiluminescence, are those which are efficient electron acceptors from the reductase and were previously shown to be those capable of catalyzing hydroxyl radical production by microsomes and the reductase. It is suggested that chemiluminescence results from (a) the direct interaction of the reduced iron chelate with the hydroperoxide (Fenton-type of reaction) to generate alkoxyl and peroxyl radicals, and (b) the generation of hydroxyl radicals, which subsequently react with the hydroperoxide to generate secondary radicals. The latter, but not the former, would be sensitive to inhibition by catalase and competitive hydroxyl radical scavengers, whereas both would be sensitive to antioxidants such as butylated hydroxytoluene. Chemiluminescence appears to be a versatile tool for studying the reductase-dependent generation of oxygen radicals and for the interaction of reductase with iron.     

Flow injection chemiluminescent determination of tetracycline using a tris(2,2'-bipyridine)ruthenium(II)-cerium(IV) sulphate system.

A flow-injection chemiluminescence method for the determination of tetracycline was developed. The method is based on an enhancement by tetracycline of the chemiluminescence light emission of tris(2,2'-bipyridine)ruthenium(II). In sulphuric acid medium, the chemiluminescence is generated by the continuous oxidation of tris(2,2'-bipyridine)ruthenium(II) by cerium (IV) sulphate. The light-emission intensity is greatly enhanced in the presence of tetracycline. Under the optimum conditions, the calibration curve is linear over the range 3.75 x 10(-8) g/mL-1.5 x 10(-5) g/mL for tetracycline with the linear equation: deltaINT = 205.898 x C - 20.442 (R2 = 0.9974). The detection limit is 3.27 x 10(-8) g/mL. The proposed method was also successfully used to determine tetracycline in pharmaceutical formulation (mean recovery of tetracycline, 100.7%).     

Chemiluminescence detection with separation techniques for bioanalytical applications

Chemiluminescence detection is known to be a sensitive, selective, and versatile method that can be used in combination with separation techniques such as high-performance liquid chromatography, capillary electrophoresis, and chip electrophoresis. This article reviews the bioanalytical applications of a combination of chemiluminescence detection and separation techniques published in the literature between 1999 and 2008. Luminol chemiluminescene, peroxyoxalate chemiluminescence, and electrochemiluminescence have been mainly used for bioanalytical application. In this paper, only the applications of the method for the analysis of biosamples, serum, plasma, urine, and tissue samples are discussed.     

Imaging of chemiluminescent reactions in mesoscale silicon-glass microstructures

Chemiluminescent reactions in mesoscale analytical structures (chips) containing micrometer-sized interconnecting channels and chambers (pL-nL total volume) were imaged. The chips were fabricated by bonding Pyrex glass to etched pieces of silicon using a high-temperature diffusive bonding technique. In initial experiments light emission from an enhanced chemiluminescent horseradish peroxidase reaction and from a peroxyoxalate reaction contained in straight channels (300 μm wide × 20μ deep; volume 70.2 nL) and open chambers (812 μm wide, 400 μm deep, 5.2 mm long) linked by channels (100μm wide, 20 μm deep) to an exit and entry port were studied using a specially modified microplate holder and an Amerlite microplate luminometer. Light emission from more complex structures (two chambers interconnected by a branching channel 100 μm wide, 20 μm deep) filled with a solution containing alkaline phosphatase, Emerald, and CSPD^TM was imaged using a Photometrics Star 1 CCD camera. Detailed investigation of the detection and spatial resolution of the signal was performed on a Berthold Luminograph LB 980 using both the enhanced chemiluminescent horseradish peroxidase reaction and a peroxyoxalate reaction. We successfully resolved light emission from silicon structures with dimensions 100 μm wide and 20 μm deep. These simple silicon structures served as models for more complex designs that will be used for simultaneous multi-analyte assays in which an imaging system resolves and quantitates light emission from different locations on a silicon-glass analytical device.     

Determination of serum glucose by horseradish peroxidase-catalysed imidazole chemiluminescence coupled to a micro-flow-injection system.

The reactivity of flow-injection (FI)-horseradish peroxidase (HRP)-catalysed imidazole chemiluminescence (CL) was studied for continuous determination of hydrogen peroxide (H(2)O(2)) and serum glucose with immobilized glucose oxidase. Light emission by the HRP-catalysed imidazole CL was obtained when immobilized HRP, alkaline imidazole (in Tricine solution, pH 9.3) and H(2)O(2) were reacted at room temperature. The optimal pH for the CL reaction was 9.3 and the optimal concentration of imidazole was 100 micromol/L. When no imidazole was added, the light intensity of the same H(2)O(2) specimen decreased to a level that could not be quantitatively determined. The spectrum of the light emitted by imidazole CL was in the range 400-600 nm with a peak at 500 nm. The calibration equation for determination of H(2)O(2) was y = 9860x(2) + 3830x + 11,700, where y = light intensity (RLU) and x = concentration of H(2)O(2) (micromol/L). The detection limit of H(2)O(2) was 5 pmol, and the reproducibility of the H(2)O(2) assay was 2.3% of the coefficient of variation (H(2)O(2) 48 micromol/L, n = 13). The CL method was successfully applied to assay glucose after on-line generation of H(2)O(2) with the immobilized glucose oxidase column, resulting in good reproducibility (CV = 3.3% and 1.0% for the standard glucose and the control serum, respectively).     

Comparison of seven bio- and chemiluminescent reagents for in situ detection of antigens and nucleic acids

Bio- and chemiluminescence have proved sensitive enough to compete with chromogenic and radioisotopic tracers for in situ detection. However, they must also provide a discriminant morphological analysis of the specific signal. We have tested seven bio-or chemiluminescent reagents for tissue antigen and nucleic acid detection by immunocytochemistry (ICC) or in situ hybridization (ISH). They were based on luminescent detection of peroxidase, aikaline phosphatase, β-galactosidase or xanthine oxidase. We also explored whether high molecular weight polymers could increase the spatial definition of the photon emission. An ICCD camera was used to collect the light signal provided by immunolabelling of endothelial cells and by ISH of human papilloma virus on cell smears. Among the enzyme-luminescent substrate combinations tested, the enhanced luminol chemiluminescence (ECL) gave the best resolution of the specific signal. The other systems were mainly hampered by a high diffusion of the reaction product over the tissue section. Unfortunately, in this case, the high molecular weight polymers tested were inefficient. However, the addition of polyvinylalcohol (PVA) or polyvinylpyrrolidone (PVP) significantly improved respectively the definition and intensity of ECL photon emission. We demonstrate that chemiluminescence gives a morphological resolution allowing histological examination. The extension of this new application, now depends on physicochemical adaptation of chemiluminescent reagents to the constraints of tissue detection.     

Microcalorimetric investigation into the metabolic activity of rat caecal flora in the presence of different sugars and sugar substitutes

When adapting young rats to different sugar substitutes (sorbitol, PolydextroseR and PalatinitR), effects were seen in the caecal morphology and caecal content e.g. bacterial concentration, which did not occur when adapting rats to sugars (glucose, sucrose). For in vitro studies, anaerobic growth of caecal flora in thioglycollate medium with and without the respective substances was monitored by continuous measurement of heat production, optical density and pH. Additionally, biochemical analyses and light microscopic observations were performed in order to detect differences between adapted and non-adapted flora. In particular the microcalorimetric data furnished valuable information about alterations in bacterial metabolic activity after adaptation to sugars and sugar substitutes, and clearly indicated that all the substances tested influenced the metabolism of caecal flora.     

Non‐enzymatic aqueous peroxyoxalate chemiluminescence immune detection using a CCD camera and a CMOS device

A new method for non‐enzymatic aqueous peroxyoxalate chemiluminescence (POCL) biomolecular detection using imaging chip‐based devices has been developed. A water‐soluble amide of oxalic acid was synthesized and used in the investigation and characterization of POCL immunodetection in an aqueous environment. Six fluorescent dyes commonly used in biological detection were tested, and the intensity of light generated from the aqueous POCL reactions was characterized in the liquid phase. Direct detection sensitivity comparisons between a standard fluorescent method and this POCL method were performed in both liquid and solid phases. Results showed that detection sensitivity using the POCL method is comparable to that of the fluorescent method. POCL biomolecular detection on a nitrocellulose membrane was also investigated using a charge‐coupled device (CCD) camera. Again, POCL detection sensitivity proved to be comparable to that using the fluorescent detection method. In an application of aqueous POCL biomolecular detection, Staphylococcus aureus enterotoxin B (SEB) and its antibody were used to demonstrate immuno‐ and affinity detection. For further applications, such as DNA and protein arrays, simultaneous detection of biomolecules labelled with different fluorescent labels was investigated, using a complementary metal oxide semiconductor (CMOS) colour imaging chip. Copyright © 2008 John Wiley & Sons, Ltd.     

Flow injection assays with chemiluminescence and bioluminescence detection — A review

This paper reviews publicaions that combine the technique of flow injection (FI) with chemiluminescence (CL) and bioluminescence (BL) detection, from the earliest papers in 1979/80 to mid-1992, and refers exclusively to reactions occurring in solution. Airsegmented systems and liquid chromatography with CL detection are not considered unless FI has been used to pre-optimize the system. The applications have been categorized in terms of the type of CL reaction; there are separate entries for luminol, peroxyoxalate, other CL reactions and BL reactions. Each of the four sections includes a table of applications that lists the analyte, the nature of the reaction, the sample matrix and the limit of detection.     

Increased NADPH-dependent chemiluminescence by microsomes after chronic ethanol consumption

The generation of reactive oxygen intermediates by microsomes from ethanol-fed rats and pair-fed controls was determined by assaying for NADPH-dependent chemiluminescence. In the absence or presence of added ferric complexes, microsomal light emission was elevated several-fold after chronic ethanol consumption. Iron complexes such as ferric-citrate or ferric-ATP stimulated, while ferric-EDTA, inhibited microsomal chemiluminescence. Freeze-thawing the microsomes to elevate their content of lipid hydroperoxides resulted in large increases in chemiluminescence; under all conditions, the light emission remained several-fold higher with microsomes from the ethanol-fed rats. Chemiluminescence was not sensitive to superoxide dismutase, catalase, or the hydroxyl radical scavenging agent, dimethyl sulfoxide, but was inhibited by antioxidants and by glutathione. Replacing air with a mixture of 50% nitrogen-50% air or 50% carbon monoxide-50% air had no effect on chemiluminescence by microsomes from the pair-fed controls. However, the chemiluminescent response by microsomes from the ethanol-fed rats was inhibited about 50% by the nitrogen mixture, and was further inhibited (about 75% of values found with 100% air, and 50% of values found with 50% nitrogen-50% air) with the carbon monoxide mixture. The sensitivity to carbon monoxide suggests the possibility that the alcohol-inducible cytochrome P-450 isozyme may contribute, in part, to the elevated light emission produced by microsomes from the ethanol-fed rats. The increase in chemiluminescence by microsomes after chronic ethanol consumption appears to reflect an elevated level of lipid hydroperoxides as well as an increased rate of generation of reactive oxygen species.     

Chemiluminescence flow-through biosensor for glucose with eggshell membrane as enzyme immobilization platform

In this study, a new chemiluminescence (CL) flow-through biosensor for glucose was developed by immobilizing glucose oxidase (GOD) and horseradish peroxidase (HRP) on the eggshell membrane with glutaraldehyde as a cross-linker. The CL detection involved enzymatic oxidation of glucose to D-gluconic acid and hydrogen peroxide (H2O2) and then H2O2 oxidizing luminol to produce CL emission in the presence of HRP. The immobilization condition (e.g., immobilization time, GOD/HRP ratio, glutaraldehyde concentration) was studied in detail. It showed good storage stability at 4 degrees C over a 5-month period. The proposed biosensor exhibited short response time, high sensitivity, easy operation, and simple sensor assembly, and the proposed biosensor was successfully applied to the determination of glucose in human serum.