Satellite high-performance liquid chromatography for trace sample cleanup.
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Chromatography, High Pressure Liquid↗
Biomedical subjects
A Abushamaa
Publications and source records attributed to A Abushamaa.
General method for determining ethylene oxide and related N7-guanine DNA adducts by gas chromatography-electron capture mass spectrometry.
A 112-micrograms sample of DNA was spiked with 103 pg of N7-(2'-hydroxyethyl)guanine and 100 pg of N7-(2'-hydroxyethyl-d4)guanine, the internal standard. The sample was subjected to the following sequence of steps: heating at 100 degrees C, precipitation of the DNA with HCl, reaction with nitrous acid to form the corresponding xanthines, reaction twice with pentafluorobenzyl bromide (first to derivatize NH, then OH), solid-phase extraction on silica and detection by gas chromatography-electron capture mass spectrometry. The absolute, overall yield of final product for both the analyte and internal standard was 9.7%. Conveniently, the three chemical reactions are conducted sequentially in the same vial and, aside from a washing step, are separated only by evaporations. Corresponding N7-guanine methyl, phenyl and styrene oxide adducts were detected at about the 50-ng level by the procedure, to indicate the generality of the method.
DNA↗
Chemical transformation/derivatization of O6-methyl- and O6-(hydroxyethyl)guanine for detection by GC-EC/MS.
In this project we set out to make an important class of DNA adducts, comprising O6-alkyl and O6-(hydroxyalkyl)guanines, susceptible to sensitive detection by GC-EC/MS. While existing literature indicated that pentafluorobenzylation would be useful for the ring NH site on these compounds, how to best overcome the polarity of the exocyclic NH2 and OH groups, without losing the O6-alkyl moiety, was less clear. Working with O6-methylguanine and O6-(2'-hydroxyethyl)guanine as representative analytes, we found that the NH2 group could be converted into fluoro without loss of the O6 substituent. For the OH group, a comparison of several derivatives (OR') led to R' = tert-butyl as the best choice at this stage. The latter work, especially via NMR, also allowed exact structural assignments to be made for the N7 and N9 pentafluorobenzyl isomeric derivatives that formed. Of these R' derivatives, the N7 isomers migrated slower on silica-TLC, had higher GC retention times, had lower responses by GC-EC/MS, and were preferentially destroyed as the GC column aged. However, the N9 isomer was slower on TLC when the OH was not derivatized. This behavior was rationalized using a concept of "polar footprint" for the derivatives. The concept also seemed to explain the puzzling GC-EC/MS behavior of some related compounds in our laboratory. Apparently the polar footprint should be minimized in designing derivatives for trace detection by GC-EC/MS.
Chromatography, Thin Layer↗