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Measurement of Hydroxyl-Radical Formation in the Rat Striatum by In Vivo Microdialysis and GC-MS

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Abstract

A GC-MS method was developed for measuring hydroxyl-radical capture products of salicylic acid, a common trapping agent for this reactive oxygen species, in samples obtained by in vivo cerebral microdialysis experiments. The assay employed liquid–liquid extraction followed by derivatization of 2,3- and 2,5-dihydroxybenzoic acid, along with 3,5-dihydroxybenzoic acid added as an internal standard. Due to their simple electron ionization mass spectra featuring [M–57]+ ions through the loss of tertiary alkyl group from the corresponding molecular ions, tert-butyldimethylsilyl (TBDMS) derivatives afforded straightforward method development based on selected-ion monitoring. In addition, tandem mass spectrometry probing collision-induced dissociation of [M–57]+ ions obtained from the isomeric tert-butyldimethylsilyl derivatives revealed characteristic differences in the resultant product-ion spectra. Our work has demonstrated the applicability of GC-MS for the assay of microdialysates for 2,3- and 2,5-dihydroxybenzoic acid by confirming that local administration of the excitotoxic glutamate into the rat striatum significantly increased in vivo hydroxyl-radical production in this brain region and that subsequent systemic administration of α-phenyl-tert-butylnitrone reversed glutamate-induced oxidative stress.

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Acknowledgments

Financial support for this work was provided in part by the grant NS044765 from the National Institute of Health (Bethesda, MD, USA), and the STARS program of the University of North Texas Health Science Center. Laszlo Prokai is the Robert A. Welch Professor of the University of North Texas Health Science Center.

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Authors and Affiliations

  1. Department of Molecular Biology and Immunology, University of North Texas Health Science Center, 3500 Camp Bowie Boulevard, Fort Worth, TX, 76107, USA

    Vien Nguyen, Darius V. Bonds & Laszlo Prokai

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  1. Vien Nguyen
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  2. Darius V. Bonds
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  3. Laszlo Prokai
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Correspondence to Laszlo Prokai.

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Nguyen, V., Bonds, D.V. & Prokai, L. Measurement of Hydroxyl-Radical Formation in the Rat Striatum by In Vivo Microdialysis and GC-MS. Chroma 68 (Suppl 1), 57–62 (2008). https://doi.org/10.1365/s10337-008-0703-6

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