A GC/MS method for the analysis of 5-methyl-2?-deoxycytidine in DNA

Randt, C.; Linscheid, Michael

doi:10.1007/bf01204103

Z. Anal. Chem.

1989

DOI: 10.1007/bf01204103

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A GC/MS method for the analysis of 5-methyl-2?-deoxycytidine in DNA

C. Randt

Michael Linscheid²

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Cited by 3 publications

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“…In 1976, a mass spectrometry technique was described to obtain higher sensitivity in the detection and identification of 5-methylated cytosine residues from intact underivatized DNA , Separation and detection of 5-methylcytosine was also obtained after derivatization with trifluoroacetic anhydride . A further study described the use of HPLC combined with GC/MS for the achievement of better specificity for the identification of the free DNA bases following derivatization with bis(trimethylsilyl) trifluoroacetamide. , The application of GC/MS to DNA hydrolyzates has been limited, however, because of the intrinsic polarity of these compounds …”

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confidence: 99%

A Method to Assess Genomic DNA Methylation Using High-Performance Liquid Chromatography/Electrospray Ionization Mass Spectrometry

et al. 2002

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Eukaryotic DNA is methylated at some cytosine residues, and this epigenetic feature performs critical functions. We developed a method for quantitative determination of 5-methyl-2'-deoxycytidine in human DNA using liquid chromatography/electrospray ionization mass spectrometry (LC/ESI-MS). The DNA was enzymatically hydrolyzed by sequential digestion with three enzymes. DNA hydrolyzates were subsequently separated by reversed-phase high-performance liquid chromatography in isocratic mode. The four major DNA bases and 5-methyl-2'-deoxycytidine were resolved and eluted in 13 min. Identification of 2'-deoxycytidine and 5-methyl-2'-deoxycytidine was obtained by combined diode array UV spectra analysis and mass spectra of chromatographic peaks. The isotopomers [15N3]-2'-deoxycytidine and (methyl-d3,ring-6-d1)-5-methyl-2'-deoxycytidine were used as internal standards. Ions of m/z 126 and 130 were used to detect 5-methyl-2'-deoxycytidine and its isotopomer, and ions of m/z 112 and 115 were used to detect 2'-deoxycytidine and its stable isotopomer, respectively. The DNA methylation status was calculated on the basis of the amount of 5-methyl-2'-deoxycytidine per microgram of DNA with percent relative standard deviations (%RSD) for a method precision of 7.1 (within-day) and 5.7 (day-to-day). This method also allows the measurement of 5-methyl-2'-deoxycytidine expressed as a percentage of total deoxycytidine residues in genomic DNA with %RSD for method precision of 1.9 (within-day) and 1.7 (day-to-day). This LC/MS method for quantitative determination of genomic DNA methylation status is rapid, sensitive, selective, and precise.

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confidence: 99%

A Method to Assess Genomic DNA Methylation Using High-Performance Liquid Chromatography/Electrospray Ionization Mass Spectrometry

et al. 2002

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A new mass spectrometric approach to detect modifications in DNA

Janning

Schräder

Linscheid

et al. 1994

Rapid Comm Mass Spectrometry

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A new approach is described for the enzymatic digestion of DNA yielding oligonucleotides ranging from dinucleoside monophosphates to octanucleoside heptaphosphates. DNA was digested by means of the benzon nuclease, an unspecific nuclease, and alkaline phosphatase to remove the terminal phosphate. The mixture of oligonucleotides was separated using capillary-zone electrophoresis with a buffer system, yielding a rather strong electro-osmotic flow. The oligomers are separated into groups with nucleotides of the same chain length. The separation capillary was used as the innermost capillary of an electrospray spraying system. Negative molecular ions of the nucleotides were recorded using a home-built interface and ion source for a sector-field mass spectrometer. This approach allows the facile detection of DNA modifications since they lead not only to differences in mass, but also can possess altered electrophoretic mobility. For modifying reactions which exhibit sequence specificity, the information is retained in the oligomers. Thus, reactions of DNA with electrophiles can be evaluated at different levels, since in longer chains, even complex sequence specificity may be apparent. Results from calf thymus DNA digests and preliminary experiments with DNA adducts with styrene oxide are discussed.

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Determination of styrene oxide adducts in DNA and DNA components

Schräder

Linscheid

1995

Journal of Chromatography A

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A GC/MS method for the analysis of 5-methyl-2?-deoxycytidine in DNA

Cited by 3 publications

References 6 publications

A Method to Assess Genomic DNA Methylation Using High-Performance Liquid Chromatography/Electrospray Ionization Mass Spectrometry

A Method to Assess Genomic DNA Methylation Using High-Performance Liquid Chromatography/Electrospray Ionization Mass Spectrometry

A new mass spectrometric approach to detect modifications in DNA

Determination of styrene oxide adducts in DNA and DNA components

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