Simultaneous HPLC-MS-MS quantification of 8-iso-PGF2α and 8,12-iso-iPF2α in CSF and brain tissue samples with on-line cleanup

Department Pathology and Laboratory Medicine, University of Pennsylvania, 3400 Spruce Street, Philadelphia, PA 19104, USA.
Journal of chromatography. B, Analytical technologies in the biomedical and life sciences (Impact Factor: 2.73). 08/2010; 878(24):2209-16. DOI: 10.1016/j.jchromb.2010.06.029
Source: PubMed


Quantitation of isoprostanes such as 8-iso-PGF(2alpha) and 8,12-iso-iPF(2alpha)-VI in biological fluids has been proposed as a reliable test of oxidant stress and inflammation in a variety of disorders. This paper presents a liquid chromatography method with tandem mass spectrometry detection for the simultaneous analysis of these two isoprostanes in human CSF and brain tissue samples. An API 5000 triple quadrupole instrument (AB Sciex, Foster City, CA, USA) with an APCI ion source was used in this study. Aliquots of CSF samples (0.25mL) were treated with a methanol:zinc sulfate mixture followed by on-line cleanup on an extraction column (Validated-C(18)) with 0.1% formic acid. The brain tissue samples were homogenized and lipids were extracted using Folch solution. Solid-phase extraction columns (C(18)) were used for the purification of the brain isoprostane fraction. Chromatographic separation was achieved using an analytical column (Synergi C(18) HydroRP) with 0.1% formic acid in water and a mixture of methanol:acetonitrile under isocratic conditions. The mass spectrometer was operated in the MRM scan and negative ion mode. The quadrupoles were set to detect the molecular ions [M-H](-) and high mass fragments of isoprostanes: m/z 353-->193amu (8-iso-PGF(2alpha)) and m/z 353-->115amu (8,12-iso-iPF(2alpha)-VI) and their deuterated internal standards: m/z 357-->197amu (8-iso-PGF(2alpha)-d(4)) and m/z 364-->115amu (8,12-iso-iPF(2alpha)-VI-d(11)). The lower limit of quantification was 2.5pg/mL for 8-iso-PGF(2alpha) and 5.0pg/mL for 8,12-iso-PF(2alpha)-VI for the CSF method and 10.0pg/0.1g of tissue and 30.0pg/0.1g of tissue for 8-iso-PGF(2alpha) and 8,12-iso-iPF(2alpha)-VI, respectively, for the brain tissue method. No ion suppression or enhancement of the detection of 8-isoPGF(2alpha), 8,12-isoPF(2alpha)-VI or both internal standards was found.

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Available from: Magdalena Korecka, Jan 14, 2015
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    • "In the past, brain tissues are usually collected from small animals like rats or mice, and the whole brain was used for isoprostanoids and isofuranoids analysis. Currently, bigger brain samples such as pig brain and post-mortem human brain (Tables I and II) are used for more detailed analysis [9] [11] [33] [43] [50]. For instance, part of the brain like frontal lobe, which is a region associated with AD can be divided for more meticulous analysis. "
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    ABSTRACT: Abstract Isoprostanoids and isofuranoids are lipid mediators that can be formed from omega-3 and omega-6 polyunsaturated fatty acids (PUFAs). F2-isoprostanes formed from arachidonic acid, especially 15-F2t-isoprostane are commonly measured in biological tissues for decades as the biomarker for oxidative stress and diseases. Recently, other forms of isoprostanoids derived from adrenic, eicosapentaenoic and docosahexaenoic acids namely F2-dihomo-isoprostanes, F3-isoprostanes and F4-neuroprostanes respectively, and isofuranoids including isofurans, dihomo-isofurans and neurofurans are reported as oxidative damage markers for different metabolisms. Blood and urine are the most widely used samples in measuring lipid peroxidation products but not limited to these; other biological fluids, specialized tissues and cells can also be determined. In this review, measurement of isoprostanoids and isofuranoids in novel biological samples by GC-MS, GC-MS/MS, LC-MS and LC-MS/MS will be discussed.
    Full-text · Article · Sep 2014 · Free Radical Research
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    • "We employed standardized scores to form a single isoprostane measure because the concentration of iso-iPF2␣-VI was approximately 3.6-fold greater than iPF2␣-III (see Korecka et al., 2010 for details) across the sample, and the use of standardized scores mitigated any potential influence of these concentration differences on the formation of the composite F2-isoprostane measure. GENLIN and follow-up t-tests, as described for iPF2␣-III and iso-iPF2␣-VI, were used to compare smokers and neversmokers on the standardized composite isoprostane level. "
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    ABSTRACT: Background Cigarette smoking in adults is associated with abnormalities in brain neurobiology. Smoking-induced central nervous system oxidative stress (OxS) is a potential mechanism associated with these abnormalities. The goal of this study was to compare cognitively-normal elders on cerebrospinal fluid (CSF) levels of F2-isoprostane biomarkers of OxS. Methods Elders with a lifetime history of smoking (smokers; n = 50; 75 ± 5 years of age; 34 ± 28 pack-years; approximately 12% were actively smoking at the time of study) were compared to never-smokers (n = 61; 76 ± 6 years of age) on CSF iPF2α-III and 8,12, iso-iPF2α-VI F2-isoprostanes levels. F2-isoprostanes levels were quantitated with HPLC-atmospheric pressure chemical ionization-tandem mass spectrometry. Associations between F2-isoprostanes levels, hippocampal volumes, and cigarette exposure measures were also evaluated. Results Smokers showed higher iPF2α-III level than never-smokers. An age x smoking status interaction was observed for 8,12, iso-iPF2α-VI, where smokers demonstrate a significantly greater concentration with increasing age than non-smokers. In smokers only, higher 8,12, iso-iPF2α-VI concentration was associated with smaller hippocampal volume, and greater iPF2α-III level was related to greater pack years. Conclusions This is the first study to demonstrate that a history of cigarette smoking in cognitively-normal elders was associated with significantly elevated CSF F2-isoprostane levels and greater age-related increases in F2-isoprostane levels, and that higher F2-isoprostane levels in smokers were related to smaller hippocampal volume. These findings provide additional novel evidence that a history of chronic smoking during adulthood is associated with adverse effects on the human brain that are potentially persistent even with extended smoking cessation.
    Full-text · Article · Sep 2014 · Drug and Alcohol Dependence
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    ABSTRACT: The Alzheimer's Disease Neuroimaging Initiative (ADNI) beginning in October 2004, is a 6-year research project that studies changes of cognition, function, brain structure and function, and biomarkers in elderly controls, subjects with mild cognitive impairment, and subjects with Alzheimer's disease (AD). A major goal is to determine and validate MRI, PET images, and cerebrospinal fluid (CSF)/blood biomarkers as predictors and outcomes for use in clinical trials of AD treatments. Structural MRI, FDG PET, C-11 Pittsburgh compound B (PIB) PET, CSF measurements of amyloid beta (Abeta) and species of tau, with clinical/cognitive measurements were performed on elderly controls, subjects with mild cognitive impairment, and subjects with AD. Structural MRI shows high rates of brain atrophy, and has high statistical power for determining treatment effects. FDG PET, C-11 Pittsburgh compound B PET, and CSF measurements of Abeta and tau were significant predictors of cognitive decline and brain atrophy. All data are available at UCLA/LONI/ADNI, without embargo. ADNI-like projects started in Australia, Europe, Japan, and Korea. ADNI provides significant new information concerning the progression of AD.
    Full-text · Article · May 2010 · Alzheimer's & dementia: the journal of the Alzheimer's Association
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