Chemiluminescence Emission Spectrum Research Articles

A novel chemiluminescent flow injection analysis of trace amounts of rutin by its inhibition of the luminol–hydrogen peroxide reaction catalyzed by tetrasulfonated colbalt phthalocyanine

Based on the inhibition effect of rutin on the luminol-hydrogen peroxide chemiluminescence (CL) system catalyzed by tetrasulfonated colbalt phthalocyanine (CoTSPc), a sensitive flow-injection CL method has been developed for the determination of rutin. The CL reaction mechanism was carefully investigated by examining CL emission spectra, UV-visible spectra and variation of reaction conditions. It was found that there existed a linear relationship between CL intensity and the concentration of rutin in the range of 8.0 x 10(-9) to 1.0 x 10(-6) mol L(-1), and the detection limit is 3.8 x 10(-9) mol L(-1). This proposed method is sensitive, convenient and simple, and has been applied to the determination of rutin in commercial rutin tablets with satisfactory results.

Interfacial-specific Chemiluminescence at Liqud/Liquid Interfaces or at O/W Emulsions Containing Lucigenin in Oil Phase

The enhanced effect of chemiluminescence (CL) of lucigenin (Luc(2+)) at a flat liquid/liquid interface and at O/W emulsions was demonstrated. The CL was emitted when Luc(2+) was transferred from an organic phase to an aqueous one containing NaOH and H(2)O(2). The CL intensity and emission spectrum depended strongly on the kind of organic solvent. We attempted to apply the CL reaction of Luc(2+) in the O/W emulsion system to the analysis of H(2)O(2). The available analytical conditions were discussed from the aspects of the kind of organic solvent, the reaction time and wavelength detected.

Chemiluminescence determination of gossypol at trace levels using flow injection as sample introduction techniques

A simple and sensitive flow injection method with chemiluminescence (CL) detection is developed for the determination of gossypol at trace levels. The method is based on the reaction of luminol with ferricyanid in a sodium hydroxide medium sensitized by gossypol. Under the optimum conditions, the CL intensity is proportional to the concentration of gossypol over the range of 1.11 × 10−17 − 2.78 × 10−16 M in an acidic solution and 8.00 × 10−11 − 7.39 × 10−8 M in a neutral solution with correlation coefficients of 0.9983 and 0.9905, respectively. The detection limit is 1.48 × 10−18 M (3σ). The proposed method has been applied to the determination of gossypol in cottonseeds and pharmaceutical preparations with satisfactory results. The CL mechanism is discussed by examining the CL emission spectrum and the effect of various free radical scavengers on the CL emission intensity.

Flow injection determination of adenine at trace level based on luminol–K 2Cr 2O 7 chemiluminescence in a micellar medium

A novel flow injection chemiluminescence (CL) analysis method for the determination of adenine in the presence of sodium dodecylbenzene sulfonate (SDBS) surfactant micelles is described. This method is based on the luminescent properties of the luminol–K 2Cr 2O 7–adenine in alkaline medium sensitized by SDBS. The optimized experimental conditions were evaluated and the possible mechanism was discussed by examining CL emission spectrum and the effect of various free radical scavengers on CL emission intensity. The CL increase is linearly related to the concentration of adenine in the range of 2.92 × 10 −6 to 4.38 × 10 −10 mol l −1 with a detection limit of 2.46 × 10 −10 mol l −1 (S/N = 3). The relative standard deviation for 2.92 × 10 −7 mol l −1 samples was 1.67% ( n = 12). The proposed method has been applied to the determination of adenine in human serum.

Enhancement in Chemiluminescence by Carbonate for Cobalt(II)-catalyzed Oxidation of Luminol with Hydrogen Peroxide

Chemiluminescence (CL) from the cobalt(II)-catalyzed oxidation of luminol with hydrogen peroxide was dramatically enhanced by the presence of carbonate. The CL signal increases by several orders of magnitude over a wide range of concentrations of Co(II), luminol, or hydrogen peroxide. A limit of detection of 10 - 1 2 M for Co(II) and luminol and 10 - 8 M for hydrogen peroxide can be achieved. The CL emission spectrum exhibits a maximum at 425 nm, indicating that the excited 3-aminophthalate is the emitting species. ESR spin-trapping experiments revealed a large increase in the production of hydroxyl and carbonate radicals by the presence of carbonate, which is responsible for the enormous CL enhancement. Uric acid, ascorbic acid, acetaminophen, and p-hydroxyphenyl acetic acid are capable of scavenging the radicals, thereby inhibiting the CL emission. The inhibition of CL intensity can be used to determine these substances at the sub-micromolar level.

Kinetic and spectral aspects in chemiluminescence system Eu(III)/HCO3−/H2O2

Chemiluminescence (CL) of the systems containing Eu2+ or HCO3− ions and hydrogen peroxide was studied. The kinetic curves and CL emission spectra of the systems were discussed. The emission spectrum of the system containing carbonates revealed two emission maxima: the first directly after initiation of the reaction by hydrogen peroxide and the second in several seconds after the first. On the basis of the characteristic bands in the CL spectrum the emitters in the system Eu2+/HCO3−/H2O2 were identified as: excited Eu3+ ions and excited products of carbonate decomposition. The emission bands for λ=∼600 and 420nm appeared in a time distance of a few ten seconds.

Reduced hemodialysis-induced oxidative stress in end-stage renal disease patients by electrolyzed reduced water

Increased oxidative stress in end-stage renal disease (ESRD) patients may oxidize macromolecules and consequently lead to cardiovascular events during chronic hemodialysis. Electrolyzed reduced water (ERW) with reactive oxygen species (ROS) scavenging ability may have a potential effect on reduction of hemodialysis-induced oxidative stress in ESRD patients. We developed a chemiluminescence emission spectrum and high-performance liquid chromatography analysis to assess the effect of ERW replacement on plasma ROS (H2O2 and HOCl) scavenging activity and oxidized lipid or protein production in ESRD patients undergoing hemodialysis. Oxidized markers, dityrosine, methylguanidine, and phosphatidylcholine hydroperoxide, and inflammatory markers, interleukin 6 (IL-6), and C-reactive protein (CRP) were determined. Although hemodialysis efficiently removes dityrosine and creatinine, hemodialysis increased oxidative stress, including phosphatidylcholine hydroperoxide, and methylguanidine. Hemodialysis reduced the plasma ROS scavenging activity, as shown by the augmented reference H2O2 and HOCl counts (Rh2o2 and Rhocl, respectively) and decreased antioxidative activity (expressed as total antioxidant status in this study). ERW administration diminished hemodialysis-enhanced Rh2o2 and Rhocl, minimized oxidized and inflammatory markers (CRP and IL-6), and partly restored total antioxidant status during 1-month treatment. This study demonstrates that hemodialysis with ERW administration may efficiently increase the H2O2- and HOCl-dependent antioxidant defense and reduce H2O2- and HOCl-induced oxidative stress.

β-Cyclodextrin sensitized chemiluminescence of hemoglobin–hydrogen peroxide–carbonate and its analytical application

The weak chemiluminescence (CL) from the reaction of hemoglobin (Hb) with hydrogen peroxide in an aqueous carbonate solution was significantly enhanced in the presence of β-cyclodextrin (β-CD). The mechanism was discussed by examining CL emission spectrum and the effect of various free radical scavengers on CL emission intensity. The β-CD could form encapsulation complex with heme, which was dissociated from Hb in alkaline medium. The formed complex showed better catalytic activity than free heme thus resulting in an enhanced CL emission intensity. A flow injection CL method for the determination of Hb ranging from 2.0×10 −9 to 1.0×10 −5 g ml −1 was developed with β-CD as a sensitizer. The detection limit (3 σ) for Hb was 1.2×10 −9 g ml −1 (1.9×10 −11 mol l −1) and the relative standard deviation (R.S.D.) for seven independent detections of 1.0×10 −7 g ml −1 Hb was 2.4%. Since metal ions could not form complex with β-CD, this method showed a good selectivity. The proposed method was used for the determination of Hb in human blood and serum.

Chemiluminescence of cobalt(II)–hydrogen peroxide–hydrogencarbonate in the absence of luminescent reagents

A novel chemiluminescence (CL) system, cobalt(II)–hydrogen peroxide–hydrogencarbonate without luminescent reagents, was described. When cobalt(II) was injected into mixed solution of 0.4 mol l −1 hydrogencarbonate and 0.01 mol l −1 hydrogen peroxide in a flow injection system, CL occurred, which was significantly enhanced when rhodamine B coexisted. The CL emission intensity was found correlation with the concentration of cobalt(II) in the range of 2.0×10 −9–1.0×10 −5 mol l −1 with a detection limit of 1.2×10 −9 mol l −1 cobalt(II). The relative standard derivative (RSD) for 5.0×10 −7 mol l −1 cobalt(II) was 1.6% based on nine repetitive measurements. In addition to cobalt(II), other metal ions were also investigated, and only chromium(III) gave out measurable CL emission. The possible mechanism was discussed based on the CL emission spectrum.

Influence of fluorescer concentration on peroxyoxalate chemiluminescence spectra

A study of the chemiluminescence emission spectra of the peroxyoxalate chemiluminescence system has been undertaken, and the results show that fluorescer concentration has a significant impact on the shape and λ max of the spectra. As fluorescer concentration increases, the shape of the spectra changes and λ max shifts gradually to longer wavelength. For the 12 substituted bis(phenylethynyl)anthracences (BPEAs) studied, a 29–38 nm bathochromic shift was generally found. Variation of oxalate, hydrogen peroxide and sodium salicylate concentration had no observable influence on the shape and λ max of the spectra.

A Bifunctional Luminogenic Substrate for Two Luminescent Enzymes: Firefly Luciferase and Horseradish Peroxidase

Horseradish peroxidase (HRP) catalyzes the oxidative chemiluminescent reaction of luminol, and firefly luciferase catalyzes the oxidation of firefly d-luciferin. Here we report a novel substrate, 5-(5′-azoluciferinyl)-2,3-dihydro-1,4-phthalazinedione (ALPDO), that can trigger the activity of HRP and firefly luciferase in solution because it contains both luminol and luciferin functionalities. It is synthesized by diazotization of luminol and its subsequent azo coupling with firefly luciferin. NMR spectral data show that the C5′ of benzothiazole in luciferin connects the diazophthalahydrazide. The electronic absorption and fluorescence properties of ALPDO are different from those of its precursor molecules. The chemiluminescence emission spectra of the conjugate substrate display biphotonic emission characteristic of azophthalatedianion and oxyluciferin. It has an optimum pH of 8.0 for maximum activity with respect to HRP as well as luciferase. At pH 8.0 the bifunctional substrate has 12 times the activity of luminol but has 7 times less activity than the firefly luciferin–luciferase system. The specific enhancement of light emission from the cyclic hydrazide part of ALPDO helped in the sensitive assay of HRP down to 2.0 × 10−13 M and of ATP to 1.0 × 10−14 mol. Addition of enhancers such as firefly luciferin and p-iodophenol (PIP) to the HRP–ALPDO–H2O2 system enhanced the light output.

Indoxyl-beta-D-glucuronide, the primary emitter of low-level chemiluminescence in plasma of hemodialysis patients

Characteristic light emission induced by the oxidation of hydroxyl radicals has been found in plasma of hemodialysis patients (Agatsuma et al., Clin Chem 1992;38:48-55). We purified a primary emitter, a chemiluminescent component peaking at 430 nm, by anion-exchange chromatography and reversed-phase HPLC. By using proton nuclear magnetic resonance and authentic indoxyl compounds, we determined the primary emitter to be indoxyl-beta-D-glucuronide. Absorption and fluorescence spectra of the purified sample coincided well with those of authentic indoxyl-beta-D-glucuronide, as did the peak in the chemiluminescence emission spectrum. Retention time of the purified sample on reversed-phase HPLC, measured by fluorescence, was also in accordance with that of indoxyl-beta-D-glucuronide. To our knowledge, this is the first identification of a primary emitter of low-level chemiluminescence from a biological source.

Weak chemiluminescence of bilirubin and its stimulation by aldehydes.

Bilirubin in an alkaline solution exhibits a weak chemiluminescence (CL) under aerobic conditions. This spontaneous CL was markedly enhanced by the addition of various aldehydes. The fluorescent emission spectrum of bilirubin, excited by weak intensity light at 350 nm, coincided with its CL emission spectrum (peak at 670 nm). CL emission from bilirubin was not quenched by active oxygen scavengers. This suggests that triplet oxygen reacts with bilirubin, and forms an oxygenated intermediate (hydroperoxide) as a primary emitter (oxidative scission of tetrapyrrole bonds in bilirubin is not involved in this CL). The Ehrlich reaction (test for monopyrroles) and hydrolsulphite reaction (test for dipyrroles) on the CL reaction mixture and unreacted bilirubin showed no differences. When the CL was initiated by singlet oxygen, rather than superoxide anion, monopyrrole, was detected in the reaction products by gel chromatography. The inhibitory effect of a scavenger of singlet oxygen on CL was eliminated in the presence of formaldehyde. Therefore, triplet carbonyl, formed by singlet oxygen through the dioxetane structure in bilirubin, is not an emitter. The reaction mechanism of bilirubin CL and the formation of a hydroperoxide intermediate is discussed in relation to the chemical structure of luciferin molecules from bioluminescent organisms.

Competitive channels in the interaction of Xe(3 P J) with Cl2, Br2 and I2. Atom transfer, excitation transfer, energy disposal and product rotational alignment

The collision of electronically excited rare-gas atoms, e.g. Xe[np5(n+ 1)s] with the molecular halogens leads both to atom transfer (generating vibrationally and electronically excited rare gas halides) and to excitation transfer (generating electronically excited molecular halogens in a range of ion-pair excimer states). Translational energy utilisation and rotational alignment in the products of the atom transfer channel have been probed using a superthermal pulsed molecular beam technique, while the vibrational energy disposal and branching ratios determined under thermal collision conditions have been reassessed through direct inversion of the resolved chemiluminescence emission spectra. The range of halogen excimer states populated through excitation transfer has been identified by appeal to the literature.

CHEMILUMINESCENCE OF BENZO[a]PYRENE‐7,8‐DIOL *

Abstract The chemiluminescence (CL) of benzo[a]pyrene‐7,8‐diol (BP‐7,8‐diol), originally observed in liver microsomal extracts, has been produced in greater than 104 higher yield by reaction with NaOCl + H2O2. The kinetics and the substrate concentration dependence for the NaOCl/H2O2 initiated CL are the same as those for Rose Bengal photosensitization of CL. In this latter singlet oxygen generating system the CL is quenched by β‐carotene at a diffusion controlled rate of 1.2 × 1010M−1 s−1. The chemically‐initiated CL emission spectrum of BP‐7,8‐diol appears to be the same as that of the microsomal CL. The long lifetime of the CL (τ= 240 s), the production of CL by 1O2 and the effects of electron donors upon the effective rate constant for decay of CL are consistent with the formation of a dioxetane intermediate. The linear dependence of CL upon substrate concentration indicates that the emission is from a monomer excited state rather than from an exciplex as suggested previously. Under the same chemical conditions the relative light yields from benzo[a]pyrene, its 12 monohydroxy derivatives and its other dihydro‐diol derivatives range from 10−2 to 10−5 of the CL from BP‐7,8‐diol. The specificity of the chemically‐initiated CL of BP‐7,8‐diol may provide a sensitive assay for this metabolite.

Single photon counting and spectroscopy of low intensity chemiluminescent reactions

With electron multiplier phototubes selected for low dark noise and with direct light coupling, chemiluminescent reactions in solution emitting as little as 200 photons s −1 will produce noise-equivalent counting rates. With this extremely sensitive system it is possible to observe the adventitious chemical production of excited states during the course of oxidase reactions in low intensity chemiluminescence as a tracer for specific metabolic reaction rates. The microsomal metabolism of carcinogenic poly cyclic aromatic hydrocarbons and the bactericidal action of phagocytic cells are accompanied by significant chemiluminescence. In the former, the chemiluminescence emission spectrum has been measured and the mechanism appears to proceed through a dioxetane intermediate to a diradical exciplex. The Luminol chemiluminescent reaction is proposed for the absolute calibration of single photon detectors and for specifying the intrinsic sensitivity of spectrometers. Procedures for these calibrations will be described.

Chemiluminescence of firefly luciferin without enzyme.

We have been able to produce chemiluminescence in firefly luciferin without utilizing the enzyme luciferase. Following the analogous mechanism of the chemiluminescence of luminol in the organic solvent dimethyl sulfoxide, we have prepared synthetically the methyl ester of luciferin, the phosphate ester of luciferin and luciferyl adenylate by condensation in dimethyl sulfoxide with Khorana's reagent, dicyclohexylcarbodiimide and diazomethane, phosphoric acid, and adenylic acid respectively. These compounds in dimethyl sulfoxide in the presence of base emit a bright chemiluminescence. Like in vitro enzymatic bioluminescence, the luciferyl adenylate. chemiluminescence emission spectrum is dependent upon pH.