Fluorescence hydrogen peroxide determination

The first detailed investigation of the reaction kinetics was reported in 1984 (68). The reaction of bis(pentachlorophenyl) oxalate [1173-75-7] (PCPO) and hydrogen peroxide cataly2ed by sodium saUcylate in chlorobenzene produced chemiluminescence from diphenylamine (DPA) as a simple time—intensity profile from which a chemiluminescence decay rate constant could be determined. These studies demonstrated a first-order dependence for both PCPO and hydrogen peroxide and a zero-order dependence on the fluorescer in accord with an earher study (9). Furthermore, the chemiluminescence quantum efficiencies Qc) are dependent on the ease of oxidation of the fluorescer, an unstable, short-hved intermediate (r = 0.5 /is) serves as the chemical activator, and such a short-hved species "is not consistent with attempts to identify a relatively stable dioxetane as the intermediate" (68). 

Amines are another important group of analytes. Mellbin and Smith [72] compared three different fluorescent reagents, dansyl chloride, 4-chloro-7-nitrobenzo-1,2,5-oxadiazole, and o-phthaldialdehyde, for derivatization of alkylamines. The dansyl tag was found to be the most effective. Hamachi et al. [73] described the application of an HPLC-POCL method for determination of a fluorescent derivative of the synthetic peptide ebiratide. Another comparative study was done by Kwakman et al. [74], where naphthalene-2,3-dialdehyde and anthracene-2,3-dial-dehyde were evaluated as precolumn labeling agents for primary amines. The anthracene-2,3-dialdehyde derivatives were not stable, especially in the presence of hydrogen peroxide, and the POCL detection of these derivatives was therefore... 

A mixture consisting of oxalic acid, carbodiimide, fluorescer, and hydrogen peroxide is known to produce strong visible light [185], Albrecht et al. used this reaction to determine oxalate in urine [118] and serum [119]. 

The most commonplace substrates in energy-transfer analytical CL methods are aryl oxalates such as to(2,4,6-trichlorophenyl) oxalate (TCPO) and z s(2,4-dinitrophenyl) oxalate (DNPO), which are oxidized with hydrogen peroxide [7, 8], In this process, which is known as the peroxyoxalate-CL (PO-CL) reaction, the fluorophore analyte is a native or derivatized fluorescent organic substance such as a polynuclear aromatic hydrocarbon, dansylamino acid, carboxylic acid, phenothiazine, or catecholamines, for example. The mechanism of the reaction between aryl oxalates and hydrogen peroxide is believed to generate dioxetane-l,2-dione, which may itself decompose to yield an excited-state species. Its interaction with a suitable fluorophore results in energy transfer to the fluorophore, and the subsequent emission can be exploited to develop analytical CL-based determinations. 

Haapakka and Kankare have studied this phenomenon and used it to determine various analytes that are active at the electrode surface [44-46], Some metal ions have been shown to catalyze ECL at oxide-covered aluminum electrodes during the reduction of hydrogen peroxide in particular. These include mercu-ry(I), mercury(II), copper(II), silver , and thallium , the latter determined to a detection limit of <10 10 M. The emission is enhanced by organic compounds that are themselves fluorescent or that form fluorescent chelates with the aluminum ion. Both salicylic acid and micelle solubilized polyaromatic hydrocarbons have been determined in this way to a limit of detection in the order of 10 8M. 

Hydrogen peroxide was determined with fmol detection limits by using rhodamine 6G and pyrimidopyrimidine derivatives as fluorescent enhancers. The method employing the latter reagent was applied to cola drinks [92], Sensitive... 

Solid-phase extraction is routinely used to clean up extracts prior to quantitation (19,42,70, 80-82). Alternatively, endogenous fluorescent artifacts in food samples can be eliminated by oxidation with potassium permanganate/hydrogen peroxide/sodium metabisulphite. Benzyl alcohol has been used to extract riboflavin selectively without the coenzymes, permitting the determination of free riboflavin. 

Wu M, Lin Z, Durkop A, Wolfbeis OS. Time-resolved enzymatic determination of glucose using a fluorescent europium probe for hydrogen peroxide. Analytical and Bioanalytical Chemistry 2004, 380, 619-626. 

Oxidation of 2-amino-3-cyanobenzodiazepinium chloride with hydrogen peroxide in the presence of peroxidase as a catalyst gave benzimidazole, which, unlike the benzodiazepine, was fluorescent, and this reaction has been applied to the determination of hydrogen peroxide by fluorimetry (80CPB2325). 

Roulier et al. reported a sensitive and specific method for the measurement of choline and hydrogen peroxide in sea-water [45]. Choline was oxidized by choline oxidase to produce betaine and H202. The latter was used with horse-radish peroxidase to oxidize hydro-xyphenyl-propionic acid to produce a fluorescent diphenol end product. The resulting fluorescence at 410 nm (excitation at 320 nm) was proportional to the amount of H202, and could thus be used to measure the amount of choline present in the sample. Only 2-dimethyl aminoethanol interfered. The method was optimized, and used to determine 0 15 nM choline in coastal sea-water. 

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