Luminol hydroperoxide determination

The first FI-CL system for lipid hydroperoxide determination, published in 1993, was based on luminol as the CL reagent and microperoxidase as the catalyst. The optimized system was used for measuring lipid hydroperoxides in native low-density lipoprotein (LDL) (Cominacini et al. 1993). The FI-CL system consisted of two pumps, an autosampler, and a chemiluminescent detector with a T-mixing coil. Samples were injected by using an autosampler and mixed with luminescent reagent (3 (iM luminol and 1 xM microperoxidase in 0.1 M carbonate buffer, pH 10). The calibration curve was obtained using linoleic acid, and a detection limit of 3 pmol linoleic acid hydroperoxide was reached. 

Luminol derivatives produce emission of light by oxidation with oxygen and hydrogen peroxide under alkaline conditions. By utilizing this reaction, peroxides such as hydrogen peroxide and lipid hydroperoxides can be determined after HPLC separation. Metal ions [e.g., iron(II), cobalt(II), etc.] catalyzing the luminol CL reaction can also be determined. 

Miyazawa T, Fujimoto K, Suzuki T, Yasuda K. Determination of phospholipid hydroperoxides using luminol chemiluminescence high-performance liquid chromatography. Oxygen Radicals in Biological Systems, Pt. C 233, 324-332, 1994. 

The CLD methods for HPLC using isoluminol (190) with microperoxidase catalysis, for determination of lipid hydroperoxides in clinical fluids, have been reviewed. Determination of phospholipids hydroperoxides by luminol (124) CL has been reviewed . A fast RP-HPLC method (retention times 1 to 2 min) for determination of hydroperoxides and other peroxide compounds includes UVD, which is not always effective, and CLD, attained on injection of luminol (124), the CL reagent (Scheme 3), hemin (75a), a catalyst, and NaOH to raise the pH of the solution. A FLD cell may act as CLD cell if the excitation source is turned off. The selectivity of CLD is of advantage over UVD in industrial analysis thus, for example, UVD of a sample from a phenol production line based on cumene oxidation (equation 13) shows peaks for cumyl hydroperoxide (27), unreacted cumene, cumyl alcohol and acetophenone, whereas CLD shows only the 27 peak. The... 

A procedure for determination of lipid hydroperoxides in human plasma is based on kinetic measurement of the CL of luminol (124) with hemin (75a) catalysis . CLD of microperoxidase-catalyzed oxidation of luminol (124) or isoluminol (190) was applied to detection and determination of amino acid hydroperoxides after exposure to UV and y-irradiation A method for determination of hydroperoxides in the phospholipids of cultured cells uses isoluminol (190) and microperoxidase as catalyst " . Simultaneous determination of phosphatidylcholine hydroperoxides and cholesteryl ester hydroperoxides in human serum is carried out by quantitative extraction of the lipids, HPLC separation by column switching and CLD using isoluminol (190) with microperoxidase catalysis . ... 

The RP-HPLC method based on the CL reaction of luminol (124) catalyzed by Co(II) (Section in.B.2.c) can be applied for determination of peroxycarboxylic acids, esters and diacyl peroxides (see examples of LOD in equation 67, Section V.B.2.c) . The biosensor prepared according to equation 70, that is effective in the determination of hydroperoxides (Section V.B.6.b), becomes deactivated after three days of operation when trying to determine t-butyl peracetate. ... 

The HPLC method with CLD described in Section V.B.2.C for determination of hydroperoxides using luminol (124) with hemin (75a) catalysis is ineffective with dialkyl peroxides, such as di-f-butyl peroxide, cumyl propyl peroxide and cumyl 3-phenylpropyl peroxide. However, for a certain set of experimental conditions, cumyl aUyl peroxide can be determined, but the sensitivity is much lower than for hydroperoxides . ... 

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