Measurement of oxidative stress parameters using liquid chromatography–tandem mass spectroscopy (LC–MS/MS)

There is increasingly intense scientific and clinical interest in oxidative stress and the many parameters used to quantify the degree of oxidative stress. However, there remain many analytical limitations to currently available assays for oxidative stress markers. Recent improvements in software, hardware, and instrumentation design have made liquid chromatography and tandem mass spectroscopy (LC–MS/MS) methods optimal choices for the determination of many oxidative stress markers. In particular, LC–MS/MS often provides the advantages of higher specificity, higher sensitivity, and the capacity to determine multiple analytes (e.g. 4–11 oxidative stress markers per LC run) when compared to other available methods, such as gas chromatography–MS, immunoassays, spectrophotometric or flourometric assays. LC–MS/MS methods are also compatible with cleanup and sample preparation methods including prior solid phase extraction or automated two dimensional LC/LC chromatography followed by MS/MS. LC–MS/MS provides three analytical filtering functions: (1) the LC column provides initial separation as each analyte elutes from the column. (2) The first MS dimension isolates ions of a particular mass-to-charge (m/z) ratio. (3) The selected precursor ion is fragmented into product ions that provide structural information about the precursor ion. Quantitation is achieved based on the abundances of the product ions. The sensitivity limits for LC–MS/MS usually lie within the range of fg–pg of analyte per LC on-column injection. In this article, the present capabilities of LC–MS/MS are briefly presented and some specific examples of the strengths of these LC–MS/MS assays are discussed. The selected examples include methods for isoprostanes, oxidized proteins and amino acids, and DNA biomarkers of oxidative stress.