Luminol peroxidase

Cysteine/glutathione Luminol-peroxidase-Cu(II) 1-50/3-50 2.5/2.8 Solutions were saturated with oxygen 62, 63... 

In this system, choline formed by acetylcholinesterase is oxidized by choline oxidase and the hydrogen peroxide produced is determined using the luminol/peroxidase CL reaction. The sensor has been used for the analysis of Paraoxon and Aldicarb pesticides, with detection limits of 0.75 pg/L and 4 pg/ L, respectively. Recoveries in the range of 81-108% in contaminated samples of soils and vegetables were obtained. 

Primary reactions that produce hydrogen peroxide have been tested14 with a luminol-peroxidase indicator reaction, Eq. 3.24 ... 

Rain Luminol/peroxidase (or Chemiluminescence 1.0x10 -0.34mgr ... 

Detecting the presence of small, even invisible, amounts of blood is routine. Physical characteristics of dried stains give minimal information, however, as dried blood can take on many hues. Many of the chemical tests for the presence of blood rely on the catalytic peroxidase activity of heme (56,57). Minute quantities of blood catalyze oxidation reactions between colorless materials, eg, phenolphthalein, luco malachite green, luminol, etc, to colored or luminescent ones. The oxidant is typically hydrogen peroxide or sodium perborate (see Automated instrumentation,hematology). 

Chemiluminescent Immunoassay. Chemiluminescence is the emission of visible light resulting from a chemical reaction. The majority of such reactions are oxidations, using oxygen or peroxides, and among the first chemicals studied for chemiluminescence were luminol (5-amino-2,3-dihydro-l,4-phthalazinedione [521-31-3]) and its derivatives (see Luminescent materials, chemiluminescence). Luminol or isoluminol can be directly linked to antibodies and used in a system with peroxidase to detect specific antigens. One of the first appHcations of this approach was for the detection of biotin (31). 

The emission yield from the horseradish peroxidase (HRP)-catalyzed luminol oxidations can be kicreased as much as a thousandfold upon addition of substituted phenols, eg, -iodophenol, -phenylphenol, or 6-hydroxybenzothiazole (119). Enhanced chemiluminescence, as this phenomenon is termed, has been the basis for several very sensitive immunometric assays that surpass the sensitivity of radioassay (120) techniques and has also been developed for detection of nucleic acid probes ia dot-slot. Southern, and Northern blot formats (121). 

Chemiluminescence and bioluminescence are also used in immunoassays to detect conventional enzyme labels (eg, alkaline phosphatase, P-galactosidase, glucose oxidase, glucose 6-phosphate dehydrogenase, horseradish peroxidase, microperoxidase, xanthine oxidase). The enhanced chemiluminescence assay for horseradish peroxidase (luminol-peroxide-4-iodophenol detection reagent) and various chemiluminescence adamantyl 1,2-dioxetane aryl phosphate substrates, eg, (11) and (15) for alkaline phosphatase labels are in routine use in immunoassay analyzers and in Western blotting kits (261—266). 

A method of detecting herbicides is proposed the photosynthetic herbicides act by binding to Photosystem II (PS II), a multiunit chlorophyll-protein complex which plays a vital role in photosynthesis. The inhibition of PS II causes a reduced photoinduced production of hydrogen peroxide, which can be measured by a chemiluminescence reaction with luminol and the enzyme horseradish peroxidase (HRP). The sensing device proposed combines the production and detection of hydrogen peroxide in a single flow assay by combining all the individual steps in a compact, portable device that utilises micro-fluidic components. 

In the reaction of luminol, hydrogen peroxide, and horse radish peroxidase 122> the chemiluminescence intensity is proportional to the square of luminol radical concentration. The lifetime of these luminol radicals was found by ESR techniques to be about 10 sec. Titration studies revealed that luminol acts as two-electron donor during the reduction of a hydrogen peroxide-horseradish peroxidase complex. The enzyme is not involved in the reaction step leading directly to light emission. This step is formulated as... 

Advances have been achieved in recent years, such as the use of CL reagents as labels to derivatize and sensitively determine analytes containing amine, carboxyl, hydroxy, thiol, and other functional groups and their application in HPLC and CE [35, 36], the synthesis and application of new acridinium esters [37], the development of enhanced CL detection of horseradish peroxidase (HRP) labels [38], the use of immobilization techniques for developing CL-based sensors [39-42], some developments of luminol-based CL in relation to its application to time-resolved or solid-surface analysis [43], and the analytical application of electrogenerated CL (ECL) [44-47], among others. 

The luminol reaction has been used for the determination of oxidizing agents such as hydrogen peroxide, for enzymes such as peroxidase and xanthine oxidase, and for metal ions such as copper or cobalt that catalyze this CL reaction [24],... 

This approach uses a kinetic sequential principle to carry out multicomponent CL-based determinations. In fact, when the half-lives of the CL reactions involved in the determination of the analytes in mixture are appreciably different, the CL intensity-versus-time curve exhibits two peaks that are separate in time (in the case of a binary mixture) this allows both analytes to be directly determined from their corresponding calibration plots. In general, commercially available chemiluminometers have been used in these determinations, so the CL reaction was initially started by addition of one or two reaction ingredients. Thus, in the analysis of binary mixtures of cysteine and gluthatione, appropriate time-resolved response curves were obtained provided that equal volumes of peroxidase and luminol were mixed and saturated with oxygen and that copper(H) and aminothiol solutions were simultaneously injected [62, 63],... 

Pesticides Immobilized choline oxidase/peroxidase, luminol-H202 CL 4 pg/L aldicarb 0.75 pg/L paraoxon 55... 

Xanthine hypoxanthine Immobilization of peroxidase or xanthine oxidase, luminol-H202 CL 2 and 5 pM 58... 

For example, peroxidase catalyzes the reaction of luminol derivatives with hydrogen peroxide and results in an increase of the CL reaction velocity and CL intensity. Therefore, intense CL can be obtained from the analyte labeled with luminol derivatives after HPLC separation, followed by reaction with peroxidase. 

As compounds exhibiting enhancing effects on CL reactions, a variety of phenols, e.g., firefly luciferin and 6-hydroxybenzothiazole derivatives [12,13], 4-iodophe-nol [14], 4-(4-hydroxyphenyl)thiazole [15], 2-(4,-hydroxy-3 -methoxy-benzyli-dene)-4-cyclopentene-l,3-dione (KIH-201) [16], and 2-(4-hydroxyphenyl)-4,5-diphenylimidazole (HDI) and 2-(4-hydroxyphenyl)-4,5-di(2-pyridyl)imidazole (HPI)[17] (Fig. 6A), and phenylboronic acid derivatives, e.g., 4-phenylylboronic acid [18], 4-iodophenylboronic acid [19], and4-[4,5-di(2-pyridyl)-l //-imidazol-2-yl]phenylboronic acid (DPPA) [20] (Fig. 6B), in the luminol/hydrogen peroxide/peroxidase system are well known. Rhodamine B and quinine are used as sensitizers in the CL-emitting reaction between cerium (IV) and thiol compounds. This CL reaction was successfully applied to the sensitive determination of various thiol drugs [21-32],... 

Figure 6 Representative (A) phenol-type and (B) phenylboronic acid-type enhancers for luminol/hydrogen peroxide/peroxidase system. KIH-201, 2-(4 -hydroxy-3 -methoxy-benzylidene)-4-cyclopentene-1,3-dione HDI, 2-(4-hydroxyphenyl)-4,5-diphenylimida-zole HPI, 2-(4-hydroxyphenyl)-4,5-di(2-pyridyl)imidazole DPPA, 4-[4,5-di(2-pyridyl)-lH-imidazol-2-yl]phenylboronic acid).

Horseradish peroxidase HRP-catalyzed Luminol- H202 Borate buffer (pH 8.5) Microchip-based CE coupled with CL detection 7-35 nM (for HRP) 107... 

Recently, two major enzyme-catalyzed chemiluminescent reactions have become popular. These use either luminol as a substrate of peroxidase or 3-(2 -spiroadamantane)-4-methoxy-4-(3"-phosphoryloxy)phenyl-1,2-dioxetane (AMPPD) as a substrate of alkaline phosphatase (ALP). 

A fiberoptic biosensor has been used for the determination of xanthine and hypoxanthine by immobilization of xanthine oxidase and peroxidase on different preactivated membranes, which were mounted onto the tip of the fiberoptic bundle [47], The hydrogen peroxide generated was measured using the luminol reaction. A linear calibration curve of the sensors occurred in the range of 1-316 nM hypoxanthine and of 3.1-316 nM xanthine, respectively, with a detection limit of 0.55 nM. 

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