Chapter 30
Oxidation of Alkylnitrosamines via the Fenton Reagent Use of Nitrosamines To Probe Oxidative Intermediates in the Fenton Reaction 1
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David A. Wink , Raymond W. Nims , Joseph E. Saavedra , Marc F. Desrosiers , and Peter C. Ford Downloaded by UNIV OF SYDNEY on September 24, 2015 | http://pubs.acs.org Publication Date: March 28, 1994 | doi: 10.1021/bk-1994-0553.ch030
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Frederick Cancer Research and Development Center, National Cancer Institute, Frederick, MD 21702 National Institute of Standards and Technology, Gaithersburg, MD 20899 University of California, Santa Barbara, CA 98106 2
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The activation of the carcinogen N-nitrosodimethylamine (NDMA) has been shown to be mediated by cytochrome P450. The biological oxidation of this nitrosamine can follow two distinct pathways, dealkylation and denitrosation. As previously postulated (1), an intermediate common to the two pathways is the alkylnitrosamino radical which is generated via hydrogen abstraction. This species can, via a rebound mechanism, form the α-hydroxynitrosamine which rapidly decomposes through the intermediacy of a diazonium salt to dealkylation products (formaldehyde, nitrogen gas, and methylating equivalents) or can rearrange to liberate NO and the Schiff's base, ultimately leading to denitrosation products (formaldehyde, nitric oxide, and monomethylamine). Heur et al. reported that the Fenton reagent was capable of oxidizing NDMA to give exclusively denitrosation products. This also was proposed to proceed via the alkylnitrosamino radical but in this case the absence of the hemoprotein pocket precludes dealkylation from occurring, resulting in denitrosation only (2). The Fenton reaction (acidic mixture of ferrous ion and peroxide) has been stated to generate hydroxyl radical as the exclusive oxidizing intermediate (3). However,
it recently has been suggested that metallo-oxo species capable of oxidation are also present (4). Using stopped-flow techniques, we demonstrated (Figure 1) that when peroxide, ferrous ion and NDMA were mixed, a transient (A) was formed at a rate 5 times faster than the rate of formation of ferric ion (5). This suggested the presence of an oxidizing intermediate that was intercepted by NDMA prior to the formation of hydroxyl radical, and this was substantiated by the oxidation of methylene blue when this redox dye was substituted for NDMA in the reaction mixture. The absorption spectrum of the transient suggested that it might be ((H O) Fe-NO) , implying that ·ΝΟ might be generated in this reaction. As the concentration of NDMA was increased, the amount of transient A also increased, +2
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This chapter not subject to U.S. copyright Published 1994 American Chemical Society
In Nitrosamines and Related N-Nitroso Compounds; Loeppky, R., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1994.
Downloaded by UNIV OF SYDNEY on September 24, 2015 | http://pubs.acs.org Publication Date: March 28, 1994 | doi: 10.1021/bk-1994-0553.ch030
WINK ET AL.
Oxidation of Alkylnitrosamines via the Fenton Reagent
40 η τ
30 -
formation ^
20 -
CO _Q Ο 10 ι I ι ι ι ι I ι ι ι ι I
0.0
0.5
1.0
1.5
time
(s)
I
2.0
0
I I
2.5
^decay
a
0.00
0.05
0.10
[H 0 ] (M) 2
2
Rgure 1. Transient A - a b s o r b a n c e changes and kinetic behavior
In Nitrosamines and Related N-Nitroso Compounds; Loeppky, R., et al.; ACS Symposium Series; American Chemical Society: Washington, DC, 1994.
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NITROSAMINES AND RELATED ΛΓ-NITROSO COMPOUNDS
but there was little or no effect on the rate of appearance or disappearance of this species. These results can be explained by invoking the reactions of Scheme 1:
-Fe* NDMA
+ 2
Fe
3
+ H Q 2
2
« ». X k.j
H
Transient A k
22
•
Fe
+ 3
2
Downloaded by UNIV OF SYDNEY on September 24, 2015 | http://pubs.acs.org Publication Date: March 28, 1994 | doi: 10.1021/bk-1994-0553.ch030
Scheme 1
A plot of 1/abSfcf vs 1/[NDMA] was linear, with -1/X-intercept = 87 mM (representing the concentration of NDMA required to generate half the maximal amount of transient A). Comparing this value with the predicted value for the hydroxyl radical (-1/X-intercept < < 5 mM) based on known rate constants (
