A simple and inexpensive preparation of perdeuterated sorbitol for use as a biomacromolecule stabilization agent in NMR studies.

Sorbitol is an excellent protein-stabilization agent, but it is typically used at high concentrations where the 1H signals can interfere with NMR data collection and analysis. Deuteration of sorbitol can ameliorate this problem; however, perdeuterated sorbitol is not commercially available. We describe a simple and inexpensive method for preparation of perdeuterated sorbitol from perdeuterated glucose. The method is described explicitly and examples are given where the use of perdeuterated sorbitol has allowed the extraction of information, from NMR spectra, that is otherwise unobtainable.

Thermospray liquid chromatography/mass spectrometry studies on mechanisms of nucleic acid alkylation by some deuterated carcinogens.

Alkylation of liver nucleic acids of rats treated with nitrosobis(2-oxopropyl)amine (BOP), nitrosomethyl(2-oxopropyl)amine (NMOP), and (methylazoxy)methane (AOM) was studied by using deuterium-labeled compounds. Six hours after treatment of each rat with 5-10 mg of compound, nucleic acids were isolated from the liver of two rats and hydrolyzed in dilute acid, and the fraction containing 7-methylguanine (7-MeG) was separated by ion-exchange high-pressure liquid chromatography. After purification by repeated chromatography, the sample was analyzed by liquid chromatography in ammonium acetate coupled with thermospray positive ion mass spectrometry. AOM containing a deuterated methyl group proximal to the oxygen-bearing nitrogen gave 7-MeG containing three deuterium atoms (169 amu), whereas AOM having the methyl group distal to oxygen deuterated produced 7-MeG without deuterium (166 amu). This indicates that methylation of nucleic acids is through oxidation of AOM to (methylazoxy)methanol and the corresponding aldehyde. BOP-d4, in which both alpha-methylenes contained deuterium, gave rise to 7-MeG containing two atoms of deuterium (168 amu). NMOP-d5, with deuterium in the N-methyl and in the alpha-methylene, gave rise only to 7-MeG containing three deuterium atoms. The methyl-d3 group must originate from the N-methyl of NMOP, probably through formation of a methyldiazonium ion. This result lends support to a mechanism involving a Baeyer-Villiger oxidation of the ketone, rather than oxidation of the N-methyl and formation of an (oxopropyl)diazonium ion, which could give rise to a methylating agent following cyclization.

Metabolism of the Z and E isomers of N-nitroso-N-methyl-(2-oxopropyl)amine by rat hepatocytes.

The metabolism of N-nitroso-N-methyl-N-(2-oxopropyl)amine was examined using freshly isolated hepatocytes from Fischer 344 rats. As determined by high performance liquid chromatography, it was found that the E isomer was preferentially metabolized when the parent mixture was used. When the two isomers were studied separately, the E isomer was efficiently metabolized in the hepatocytic system, whereas the Z isomer was not. The kinetics of disappearance of the Z isomer during metabolism was identical to that for the reequilibration of the Z isomer to the mixture of isomers in the absence of a metabolizing system.