Implementation of HPLC methodology for the quantification of malondialdehyde in cell suspensions and liver

Soares, M. Elisa; Carvalho, Márcia; Remião, Fernando; Carvalho, Félix; Bastos, Maria de Lourdes

http://hdl.handle.net/10284/10004

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Authors

Bastos, Maria de Lourdes

Abstract(s)

Determination of malondialdehyde (MDA) is commonly used for the evaluation of lipid peroxidation in biological human samples, especially in plasma, serum, and urine, as a non‐invasive biomarker of some oxidative‐stress‐related diseases. The objective of the present study was to implement a HPLC methodology for the quantification of MDA in cell suspensions and liver, after complexing this compound with thiobarbituric acid (TBA), giving rise to the formation of the complex MDA–(TBA)2. The implemented methodology was validated and allowed a reliable separation and quantification of MDA, with high precision and recovery, and a low detection limit. The stability of MDA in the biological samples, as well as of the MDA–(TBA)2 complex, was also evaluated during storage at different temperatures and for different time periods. The data obtained from the stability studies concerning the MDA–(TBA)2 complex prepared from hepatocyte suspensions and liver samples, indicated that the chromatographic analysis should be performed within the period of 4 hr when it is maintained at ambient temperature or at 40°C, and within the period of 48 hr if the temperatures of the storage are 4°C, −20°C, or −80°C. When the biological samples are conserved for posterior preparation of the complex, MDA can be measured in the cell suspensions conserved at any of these temperatures in the subsequent 24 hr, but liver samples must be conserved at 4°C or −20°C in order to obtain reliable results of MDA.