Determination of 8-Hydroxydeoxyguanosine in Biological Tissue by Liquid Chromatography/Electrospray Ionization-Mass Spectrometry/Mass Spectrometry

Serrano, Jose A.; Palmeira, Carlos M.; Wallace, Kendall B.; Kuehl, Douglas W.

doi:10.1002/(sici)1097-0231(199611)10:14<1789::aid-rcm752>3.0.co;2-6

Cited by 67 publications

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“…In this respect, we report the simultaneous measurement of four thymine dimeric photoproducts by HPLC associated with electrospray ionization-MS/MS operating in the multiple reaction monitoring mode. The latter technique has already been applied to the sensitive detection of 8-oxo-7,8-dihydro-2Ј-deoxyguanosine, a major DNA oxidation product (43,44). The limit of sensitivity of the assay for UV-induced thymine lesions ranges between 10 and 50 fmol, depending on the photoproduct.…”

Section: Discussionmentioning

confidence: 99%

Formation of the Main UV-induced Thymine Dimeric Lesions within Isolated and Cellular DNA as Measured by High Performance Liquid Chromatography-Tandem Mass Spectrometry

Douki¹,

Court²,

Sauvaigo³

et al. 2000

Journal of Biological Chemistry

227

194

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UVB radiation-induced formation of dimeric photoproducts at bipyrimidine sites within DNA has been unambiguously associated with the lethal and mutagenic properties of sunlight. The main lesions include the cyclobutane pyrimidine dimers and the pyrimidine (6-4) pyrimidone adducts. The latter compounds have been shown in model systems to be converted into their Dewar valence isomers upon exposure to UVB light. A new direct assay, based on the use of liquid chromatography coupled to tandem mass spectrometry, is now available to simultaneously detect each of the thymine photoproducts. It was applied to the determination of the yields of formation of the thymine lesions within both isolated and cellular DNA exposed to either UVC or UVB radiation. The cis-syn cyclobutane thymine dimer was found to be the major photoproduct within cellular DNA, whereas the related (6-4) adduct was produced in an approximately 8-fold lower yield. Interestingly, the corresponding Dewar valence isomer could not be detected upon exposure of human cells to biologically relevant doses of UVB radiation.Ultraviolet radiation represents the most deleterious part of solar light to cells and has been associated with the occurrence of skin cancer (1). UVB (290 -320 nm) radiation is highly mutagenic (2, 3) and mostly induces mutations at bipyrimidine sites in cellular DNA (4 -7). Interestingly, a similar mutation spectrum was observed in the p53 gene of skin tumors cells (8,9). Altogether, these data outlined the biological role of dimeric photoproducts of pyrimidine DNA bases. In the last four decades, major efforts have been devoted to the isolation and the characterization of bipyrimidine photolesions in model compounds and isolated DNA (for reviews, see Refs. 10 and 11). These include the cis-syn and trans-syn diastereoisomers of cyclobutane dimers and the pyrimidine (6-4) pyrimidone adducts (Fig. 1). The latter lesions have been shown to undergo an efficient photoconversion into their Dewar valence isomers upon exposure to UVB light (12).Information on the rate of formation of each specific photoproduct is still needed. Indeed, most of the assays developed for the detection of UV-induced photoproducts within DNA involve the use of indirect methods that do not allow differentiation, for a given class of photoproduct, of the lesions arising from the different possible bipyrimidinic sequences. Cyclobutane pyrimidine dimers have been extensively detected by using T4 endonuclease V, which exhibits a N-glycosylase activity at the 5Ј-extremity of the lesion (13). This leads to the formation of strand breaks, which can be quantified using electrophoretic techniques. A second DNA repair system, namely the Escherichia coli Uvr ABC complex, has been used in combination with a photolyase for the quantitation of cyclobutane dimers and (6-4) photoproducts (14). Another widely applied biochemical approach involves the use of either polyclonal or monoclonal antibodies able to recognize specific classes of pyrimidine photoproducts (15)(16)(17)(18)(19)(20)(21)...

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Section: Discussionmentioning

confidence: 99%

Formation of the Main UV-induced Thymine Dimeric Lesions within Isolated and Cellular DNA as Measured by High Performance Liquid Chromatography-Tandem Mass Spectrometry

Douki¹,

Court²,

Sauvaigo³

et al. 2000

Journal of Biological Chemistry

227

194

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“…27, 28 However, 8-oxodG may form during sample pretreatment in some protocols, 19,29,30 and further oxidation may destroy 8-oxodG. 31,32 Oxidative DNA lesions can be measured without hydrolysis by using specific antibodies along with Comet assays or enzyme-linked immunosorbent assay (ELISA) 33-36 .…”

Section: Introductionmentioning

confidence: 99%

Voltammetric Microwell Array for Oxidized Guanosine in Intact ds-DNA

et al. 2013

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Oxidative stress in humans causes damage to biomolecules by generating reactive oxygen species (ROS). DNA can be oxidatively damaged by ROS, which may lead to carcinogenesis. Here we report a microfluidic electrochemical array designed to rapidly detect oxidation in intact DNA in replicate measurements. Sensor arrays were fabricated by wet-chemistry patterning of gold compact discs. The 8-sensor array is incorporated into a 60 µL microfluidic channel connected to a pump and sample valve. The array features 7 nm thick osmium bipyridyl poly(vinylpyridine) chloride [Os(bpy)2(PVP)10Cl]+ films assembled layer-by-layer with polyions onto the gold sensors. 7,8-dihydro-8-oxoguanine (8-oxodG) in ds-DNA is selectively oxidized by [Os(bpy)2(PVP)10Cl]+ in intact ds-DNA to provide catalytic square wave voltammograms (SWV). The device is easy-to-use, fast, inexpensive, reusable and can detect one 8-oxodG per 6600 nucleobases. The mass detection limit is 150-fold lower than a previously reported dip-and-read voltammetric sensor for oxidized DNA. Fast assays (<1 min) and moderate sample consumption (15 pmol DNA) suggest potential for research and clinical applications. Practical use is illustrated by detecting DNA oxidation from cigarette smoke and ash extracts in dispersions with NADPH and Cu2+.

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“…Among these methods, high performance liquid chromatography-electrochemical detection and gas chromatography (GC)-mass spectrometry (MS) are two prominent contributions in the field [8][9][10]. In addition, 32 P-postlabeling assays [11], high-performance liquid chromatography tandem mass spectrometry [12], antibody-based immunoassay [13], photoelectrochemical-based detection [14,15] and single-cell gel electrophoresis [16] have been used to determine a variety of oxidative DNA lesions. These methods have their own advantages but limitations still exist.…”

Section: Introductionmentioning

confidence: 99%

Selective enzymatic cleavage and labeling for sensitive capillary electrophoresis laser-induced fluorescence analysis of oxidized DNA bases

Wang

2015

Journal of Chromatography A

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a b s t r a c tOxidatively generated DNA damage is considered to be a significant contributing factor to cancer, aging, and age-related human diseases. It is important to detect oxidatively generated DNA damage to understand and clinically diagnosis diseases caused by oxidative damage. In this study, using selective enzymatic cleavage and quantum dot (QD) labeling, we developed a novel capillary electrophoresislaser induced fluorescence method for the sensitive detection of oxidized DNA bases. First, oxidized DNA bases are recognized and removed by one DNA base excision repair glycosylase, leaving apurinic and apyrimidinic sites (AP sites) at the oxidized positions. The AP sites are further excised by the AP nicking activity of the chosen glycosylase, generating a nucleotide gap with 5 -and 3 -phosphate groups. After dephosphorylation with one alkaline phosphatase, a biotinylated ddNTP is introduced into the nucleotide space within the DNA strand by DNA polymerase I. The biotin-tagged DNA is further labeled with a QD-streptavidin conjugate via non-covalent interactions. The DNA-bound QD is well-separated from excess DNA-unbound QD by highly efficient capillary electrophoresis and is sensitively detected by online coupled laser-induced fluorescence analysis. Using this method, we can assess the trace levels of oxidized DNA bases induced by the Fenton reaction and UV irradiation. Interestingly, the use of the formamidopyrimidine glycosylase (FPG) protein and endonuclease VIII enables the detection of oxidized purine and pyrimidine bases, respectively. Using the synthesized standard DNA, the approach has low limits of detection of 1.1 × 10 −19 mol in mass and 2.9 pM in concentration.

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Determination of 8-Hydroxydeoxyguanosine in Biological Tissue by Liquid Chromatography/Electrospray Ionization-Mass Spectrometry/Mass Spectrometry

Cited by 67 publications

References 0 publications

Formation of the Main UV-induced Thymine Dimeric Lesions within Isolated and Cellular DNA as Measured by High Performance Liquid Chromatography-Tandem Mass Spectrometry

Formation of the Main UV-induced Thymine Dimeric Lesions within Isolated and Cellular DNA as Measured by High Performance Liquid Chromatography-Tandem Mass Spectrometry

Voltammetric Microwell Array for Oxidized Guanosine in Intact ds-DNA

Selective enzymatic cleavage and labeling for sensitive capillary electrophoresis laser-induced fluorescence analysis of oxidized DNA bases

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