Chemiluminescence detection reaction process

Therefore, this sensor integrates a biochemical and a chemical reaction with a prior separation (dialysis) and chemiluminescence detection. The process involves the following steps (a) dialysis of the enzyme (6) enzymatic oxidation of the reagent (c) derivatization of hydrogen peroxide and d) detection of the chemiluminescence produced. Such an original approach offers several advantages over similar methodologies, namely ... 

According to the review by Marques et al.,75 the most frequent pretreatment used for chromium speciation is complex formation. Extraction processes are frequently used after complex formation to extract the complexes formed prior to UV-VIS detection. Recently, the simultaneous determination of Cr(III) and Cr(VI) using an in-capillary reaction, CE separation, and chemiluminescence detection was reported with LODs (Table 7.2) for Cr(III) and Cr(VI) of 0.6 and 8 pM, respectively.32... 

Before considering detector characteristics and some recent developments in chemiluminescence detection, it should be noted that analytical applications of chemiluminescence involve two types of chemiluminescent response. In the first type, the chemiluminescent molecule is used as a detection label and is, therefore, present in limiting concentration relative to the reagents used to initiate the chemiluminescent reaction. The chemical reaction will therefore be pseudo first order. The slowest process in the sequence of events leading to light emission is the reaction itself, e.g., hydrolysis, bond-breaking, and rearrangements. From Eq. 

The reaction process is considered to be the same for similar compounds, but the sensitivity of chemiluminescence has been suggested to be structure-dependent. The reactivity of a compound contributes to its sensitivity. Phenacylalcohol derivatives have been detected with different sensitivities in chemiluminescence analysis.In radical reactions, the reaction proceeds as follows in buffered solutions, a compound such as phenacylalcohol is easily attacked by oxidation if traces of a copper or iron salt are present, and the superoxide reacts with luminol or lueigenin to produce chemiluminescence as shown in Figure 11.7. 

Numerous applications involve coupling liquid-phase chemiluminescence detection to physical or chemical separation processes. Conversely, adequate selectivity can also be achieved for particular analytes in a range of sample matrices through a judicious selection of reagent and reaction conditions. Successful detection strategies have been employed for HPLC, flow analysis, electrophoresis, immunoassay labels, DNA probes, and enzyme reactions. 

An important reaction in chemiluminescence detection is the cerium(IV)—rhodamine reaction in sulfuric acid medium. In the presence of parabens, this process is enhanced, as the cerium(IV) is reduced to cerium(III) and the energy is then transferred from cerium(III) to rhodamine 6G to form the excited rhodamine 6G which emits radiation at 556 nm (Table 10.1). 

Nickel Carbonyl The extremely toxic gas nickel carbonyl can be detected at 0.01 ppb by measuring its chemiluminescent reaction with ozone in the presence of carbon monoxide. The reaction produces excited nickel(II) oxide by a chain process which generates many photons from each pollutant molecule to permit high sensitivity (315). 

Chemiluminescence. Chemiluminescence (262—265) is the emission of light duting an exothermic chemical reaction, generaUy as fluorescence. It often occurs ia oxidation processes, and enzyme-mediated bioluminescence has important analytical appHcations (241,262). Chemiluminescence analysis is highly specific and can reach ppb detection limits with relatively simple iastmmentation. Nitric oxide has been so analyzed from reaction with ozone (266—268), and ozone can be detected by the emission at 585 nm from reaction with ethylene. 

Though we and others (27-29) have demonstrated the utility and the improved sensitivity of the peroxyoxalate chemiluminescence method for analyte detection in RP-HPLC separations for appropriate substrates, a substantial area for Improvement and refinement of the technique remains. We have shown that the reactions of hydrogen peroxide and oxalate esters yield a very complex array of reactive intermediates, some of which activate the fluorophor to its fluorescent state. The mechanism for the ester reaction as well as the process for conversion of the chemical potential energy into electronic (excited state) energy remain to be detailed. Finally, the refinement of the technique for routine application of this sensitive method, including the optimization of the effi-ciencies for each of the contributing factors, is currently a major effort in the Center for Bioanalytical Research. 

Because process mixtures are complex, specialized detectors may substitute for separation efficiency. One specialized detector is the array amperometric detector, which allows selective detection of electrochemically active compounds.23 Electrochemical array detectors are discussed in greater detail in Chapter 5. Many pharmaceutical compounds are chiral, so a detector capable of determining optical purity would be extremely useful in monitoring synthetic reactions. A double-beam circular dichroism detector using a laser as the source was used for the selective detection of chiral cobalt compounds.24 The double-beam, single-source construction reduces the limitations of flicker noise. Chemiluminescence of an ozonized mixture was used as the principle for a sulfur-selective detector used to analyze pesticides, proteins, and blood thiols from rat plasma.25 Chemiluminescence using bis (2,4, 6-trichlorophenyl) oxalate was used for the selective detection of catalytically reduced nitrated polycyclic aromatic hydrocarbons from diesel exhaust.26... 

The dismutation (disproportioning) of two free radicals is accompanied by release of a portion of reaction energy as a light quantum. As the quantum yield of such a process is extremely low, the detection of this type of chemiluminescence is technically complicated. Several compounds like lucigenin and luminol have a high quantum yield after reaction with peroxide radicals. Therefore, they are widely used for the detection of these radicals, particularly in the examination of phagocyting cells. 

The acridinium ester (AE) in an AE-labeled cDNA probe hybridized to target DNA is less likely to be hydrolyzed than in the unhybridized conformation (Fig. 10) [9-11]. Single-base mismatches in the duplex adjacent to the site of AE attachment disrupt this protection, resulting in rapid AE hydrolysis [11]. Hydrolysis by a weak base renders AE permanently nonchemiluminescent. After hydrolysis, it is possible to use the remaining chemiluminescence as a direct measure of the amount of hybrid present. This selective degradation process is a highly specific chemical hydrolysis reaction, which is sensitive to the local environment of the acridinium ester. The matched duplex can be detected and quantified readily, whereas the mismatched duplex produces a minimal signal. 

Bromine has also been suggested for an indirect detection process for the determination of tetracyclines in pharmaceutical formulations [155]. A bromine/hydrogen peroxide-based electrogenerated chemiluminescence reaction is shown to be enhanced by tetracycline derivatives and detection levels down to the pg dm level are reported on the basis of this enhancement effect. 

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