Kinetic analysis of lipid-hydroperoxides in plasma.
We increased the precision of chemiluminescent procedure for measuring lipid hydroperoxides in plasma or lipoproteins by (i) escaping from extraction and chromatography of lipids, (ii) using detergent dispersed lipids, and (iii) calculating the results by fitting the photon emission rate with the integrated equation, which describes the model of the series of reactions. The use of kinetics instead of the crude integration of cps increases precision because at each measurement the correct reaction pathway is tested. This was relevant for the optimization of the analytical procedure, contributing to the elimination of possible side reactions. The relationship between lipid hydroperoxide content in the sample and cps is not linear; thus, the calculation of results through internal calibration is carried out using an exponential equation. This is in agreement with the reaction mechanism and raises the point of the linear calibration previously reported in other chemiluminescent procedures. Although sensitive and precise, this procedure suffers for being time consuming, requiring approximately 30 min per sample. Moreover, since no chromatography is used, information about the hydroperoxides in different lipid classes is missing. Obviously this will be solved when a validated procedure for quantitatively extracting lipid hydroperoxides is available.
Pastorino
A M
AM
Department of Biological Chemistry, University of Padova, Padova, Italy.
Maiorino
M
M
Ursini
F
F
D002138:Calibration; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D006801:Humans; D007700:Kinetics; D008054:Lipid Peroxides / Q000097:blood*; D008163:Luminescent Measurements; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D008054:Lipid Peroxides