Effects of Analytical Procedures on the Repeatability of Malondialdehyde Determinations in Biological Samples

Background: Malondialdehyde (MDA) is a commonly used biomarker of oxidative stress in clinical studies and has been measured in many pathological conditions during last decades. Different analytical methods have been reported for determination of MDA in biological samples in which MDA was adducted with thiobarbituric acid (TBA) to produce more sensitive chromophore and also convert it to a fluorescent compound. In spite of the routine applications of this derivatization and subsequent analysis of MDA in biomedical studies, its reliability, repeatability and reproducibility is questionable. The aim of this work is to investigate the effects of some factors on the repeatability of MDA determinations in standard solutions and also in plasma samples using spectroscopic method. Methods: MDA-TBA adduct is prepared in standard solutions and the effects of pH, temperature, reaction time, open, closed and reflux systems and the ratio of MDA and TBA is investigated by measuring the absorbance of the solution at 532 nm. These effects are also investigated in human plasma samples. Results: The best results are obtained at pH 2.5, temperature of 70 °C, reaction time of 150 minutes, reflux system and ratio of 2. Conclusion: Using the optimized conditions are resulted in better repeatability.