Quantification of 1,N6-Etheno-2′-deoxyadenosine in human urine by column-switching LC/APCI-MS/MS

Hillestrøm, Peter René; Hoberg, Anne-Mette; Weimann, Allan; Poulsen, Henrik E.

doi:10.1016/j.freeradbiomed.2004.02.068

Cited by 31 publications

(47 citation statements)

References 27 publications

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“…The corresponding levels of urinary excretion rates in the six subjects were in the ranges 14.2-117.3 pmol Ade/24 h, 53.2-117.8 pmol dC/24 h, and 9.8 -23.4 pmol dA/24 h. The basal levels here are comparable with those of dA [7,16] and dC [11] found previously. Slightly higher excretion levels of Ade than our finding have been reported by Chen and coworkers [9,12].…”

Section: Resultssupporting

confidence: 87%

“…Solid-phase extraction was performed as described previously with minor modifications [16]. The frozen urine was thawed and 2 mL aliquots were diluted 1:2 in 5 mM ammonium acetate (pH 5) and heated to 37°C for 5 min.…”

Section: Sample Handlingmentioning

confidence: 99%

“…Recently, LC-MS have been employed in determination of etheno adducts from human urine using electrospray ionization (ESI) [12][13][14]. Atmospheric pressure chemical ionization (APCI) has been used for analysis of dA and 1,N 2 -dG in vitro [15] and of dA in human urine [16].In this paper, we report that solid-phase extraction (SPE) for preconcentration combined with HPLC column switching tandem mass spectrometry can quantify urinary etheno-DNA modifications. This approach demonstrates progress in the simultaneous analysis of urinary Ade, dC, and dA in terms of specific identification and the stringent sensitivity requirements needed for low-level analysis.…”

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Quantification of urinary etheno-DNA adducts by column-switching LC/APCI-MS/MS

Hillestrøm

Weimann

Poulsen

2006

J. Am. Soc. Mass Spectrom.

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Lipid peroxidation induced etheno-DNA adducts are promutagenic and have been suggested to play a causal role in the development of human cancers. Therefore, human biomonitoring of etheno-DNA adducts in urine has been suggested as a potential marker for oxidative stress-related DNA damage. For quantitative determination, a column-switching LC/APCI-MS/MS method was developed for simultaneous analysis of Ade, dC, and dA in human urine. Quantitative validation parameters (precision, within-day repeatability, and betweenday reproducibility) yielded satisfactory results below 10%. Limit of quantification for Ade, dC, and dA was 5.3 fmol, 7.5 fmol, and 1.3 fmol on column, respectively. Mean urinary excretion rates of a six healthy volunteers were 45.8 pmol Ade/24 h, 96.8 pmol dC/24 h, and 18.1 pmol dA/24 h. The demonstrated levels of performance suggest a future applicability of this method to studies of cancer and other diseases related to oxidative stress in humans. To our knowledge, this is the first method described that allows simultaneous determination of Ade, dC, and dA in human urine samples. [2], are also generated endogenously by reactions of DNA with products derived from lipid peroxidation and oxidative stress [3]. These DNA adducts have miscoding potential and specific repair pathways supporting the hypothesis that -DNA adducts play a causal relationship in carcinogenesis [4]. Furthermore, several studies have found elevated -DNA adduct levels in disorders known as risk factors for cancers [5,6]. Therefore -DNA adducts have been proposed as biomarkers for human cancers associated with certain lifestyles or chronic inflammations [4]. Upon DNA repair, -DNA adducts are excreted in the urine and could, in that respect, be used to investigate the body burden of DNA damage.The analytical procedures for urinary DNA adducts are challenging indeed. The presence of many interfering substances in concentrations far greater than those of the DNA adducts demands high selectivity and low limit of quantitation for unequivocal identification and quantification. Methods using high-performance liquid chromatography (HPLC) with fluorescence detection have been used for dA determination [4,7] after immunoaffinity clean-up and a very tedious sample preparation. Furthermore, the internal standard was measured separately. Several etheno-DNA adducts have been determined in urine from healthy humans using gas chromatography-electron capture negative chemical ionization mass spectrometry (GC/EC-NCI/ MS) [8 -11]. Gas chromatography offers the advantage of greater peak resolution compared to conventional LC. However, the requirement for derivatization before analysis is a major limitation. Recently, LC-MS have been employed in determination of etheno adducts from human urine using electrospray ionization (ESI) [12][13][14]. Atmospheric pressure chemical ionization (APCI) has been used for analysis of dA and 1,N 2 -dG in vitro [15] and of dA in human urine [16].In this paper, we report that solid-phase extraction (SPE) for preconcent...

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Section: Resultssupporting

confidence: 87%

Section: Sample Handlingmentioning

confidence: 99%

mentioning

confidence: 99%

mentioning

confidence: 99%

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Quantification of urinary etheno-DNA adducts by column-switching LC/APCI-MS/MS

Hillestrøm

Weimann

Poulsen

2006

J. Am. Soc. Mass Spectrom.

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“…APCI also allows for the coupling of LC operating at higher flow rates compared with those used for ESI and is useful for analysing less polar compounds (85). Recently, using APCI the detection of etheno-2 0 -deoxyadenosine and malondialdehyde-2 0 -deoxyguanosine adducts was reported in human urine (86,87).…”

Section: Mass Spectrometry In the Analysis Of Dna Adductsmentioning

confidence: 99%