Environmental Health Perspectives Volume
102, Supplement 6, October 1994
[Citation
in PubMed] [Related
Articles]
Reactions of Oxidatively Activated Arylamines with Thiols: Reaction
Mechanisms and Biologic Implications. An Overview
Peter Eyer
Walther-Straub-Institut für Pharmakologie und Toxikologie, Ludwig-Maximilians-Universität
München, München, Germany
Abstract
Aromatic amines belong to a group of compounds that exert their toxic
effects usually after oxidative biotransformation, primarily in the liver.
In addition, aromatic amines also undergo extrahepatic activation to yield
free arylaminyl radicals. The reactive intermediates are potential promutagens
and procarcinogens, and responsible for target tissue toxicity. Since thiols
react with these intermediates at high rates, it is of interest to know
the underlying reaction mechanisms and the toxicologic implications. Phenoxyl
radicals from aminophenols and aminyl radicals from phenylenediamines quickly
disproportionate to quinone imines and quinone diimines. Depending on the
structure, Michael addition or reduction reactions with thiols may prevail.
Products of sequential oxidation/addition reactions (e.g., S-conjugates
of aminophenols) are occasionally more toxic than the parent compounds because
of their higher autoxidizability and their accumulation in the kidney. Even
after covalent binding of quinone imines to protein SH groups, the resulting
thioethers are able to autoxidize. The quinoid thioethers can then cross-link
the protein by addition to neighboring nucleophiles. The reactions of nitrosoarenes
with thiols yield a so-called "semimercaptal" from which various
branching reactions detach, depending on substituents. Compounds with strong
pi-donors, like 4-nitrosophenetol, give a resonance-stabilized N-(thiol-S-yl)-arylamine
cation that may lead to bicyclic products, thioethers, and DNA adducts.
Examples of toxicologic implications of the interactions of nitroso compounds
with thiols are given for nitrosoimidazoles, heterocyclic nitroso compounds
from protein pyrolysates, and nitrosoarenes. These data indicate that interactions
of activated arylamines with thiols may not be regarded exclusively as detoxication
reactions. -- Environ Health Perspect 102(Suppl 6):123-132 (1994)
Key words: aminophenols, arylamines, C-nitroso aromatics,
mutagenicity, quinone imines, radicals, reaction mechanism, thioethers,
thiols, toxicity
This paper was presented at the Fifth International Conference
on Carcinogenic and Mutagenic N-Substituted Aryl Compounds held
18-21 October 1992 in Würzburg, Germany.
This study was supported by the Deutsche Forschungsgemeinschaft.
The author thanks D. Gallemann for helpful comments.
Address correspondence to the author at Walther Straub-Institut
für Pharmakologie und Toxikologie, Ludwig - Maximilians - Universität
München, Nußbaumstraße 26 D-8000 München 2, Germany.
Telephone 89 51 45 2281. Fax 89 51 45 2224.
Introduction
Aromatic amines belong to a group of chemicals that exert their toxic
effects usually after oxidative biotransformation, primarily in the liver.
As a result, C-oxygenated and N-oxygenated products are generated,
which can be further activated to give electrophilic intermediates responsible
for toxic, allergic, mutagenic, and carcinogenic effects.
The group of aminophenols can be activated in metabolically competent
cells to give ultimate reactive intermediates like phenoxyl radicals and
quinone imines. The phenylhydroxylamines are activated by O-esterification
(glucuronidation, sulfatation, acetylation) to yield carbonium-nitrenium
ions as ultimate reactive compounds that alkylate nucleic acids under preferable
formation of N(deoxyguanosine-8-yl)arylamine adducts (1).
In addition, oxidative activation of the phenylhydroxylamines produces phenylhydronitroxide
radicals which react with cellular sulfhydryls to give thiyl radicals of
glutathione (GSH) and protein sulfhydryl groups (2). The nitrosoarenes,
formed by either 2-electron oxidation of the phenylhydroxylamines, by 1-electron
oxidation of the phenylhydronitroxide radicals, by disproportionation of
the latter, or by reduction of nitro aromatic compounds, exert their toxic
effects mainly through their thiol reactivity (3).
Last, aromatic amines also undergo extrahepatic activation to yield free
arylaminyl radicals (4,5). This kind of activation received more
attention when it was shown that prostaglandin hydroperoxidase mediated
the arachidonic acid-dependent cooxidation of arylamines to DNA-bound products
(6 ).
Since thiols usually react at high rates with all the aforementioned
electrophilic intermediates, it is of interest to know the underlying reaction
mechanisms and the toxicologic implications. This article is intended to
focus not only on detoxication reactions, as usually supposed to emergency
reactions, particularly of GSH. Evidence is growing that thiol interaction
with electrophilic intermediates may produce sufficiently stable transport
forms that can cause toxic effects in remote, sensitive target organs.
Mechanisms of Thiol Reactions with Radicals and Quinoid
Compounds
Aminyl Radicals
During our investigations on ferrihemoglobin formation by p-phenylenediamines
in human red cells, we detected formation of glutathione adducts. The same
adducts were formed when the chemically prepared radical cations were allowed
to react with GSH. In addition, some GSSG was formed. Since N-centered
free radicals of aromatic amines tend to disproportionate at high rates
(7), it was difficult to decide whether redox and addition reactions
with GSH had originated directly from the radicals or from the quinone diimines.
To reveal unambiguously the underlying microscopic events, we examined
reactions of the stable N-alkyl-p-phenylene- diamine radical
cations (Wurster's dyes) with GSH. Of these the N,N-dimethyl-p-phenylenediamine
radical cation (DMPD*+) turned out to be particularly suited
for our investigations, because its disproportionation equilibrium is near
unity at physiologic pH.
2 DMPD*+ 
DMPD +
DMQDI+ + H+ [1]
Figure 1 shows the influence of pH and additional DMPD (10-fold excess)
on the disproportionation equilibrium of DMPD*+ (8). It
is obvious that addition of the inactive DMPD increases the radical steady-state
concentration 1.8-fold and decreases the quinone diimine concentration to
1/5. By this measure, one can easily decide whether the radical or the quinone
diimine (DMQDI+) is the ultimate reactive compound (Table 1).
Figure 1. Influence
of pH and additional DMPD on the disproportionation equilibrium of DMPD*+.
The actual radical proportion was determined spectroscopically without and
with a 10-fold excess of DMPD. Adapted from Störle and Eyer (8).
From the kinetics (stopped-flow spectroscopy) of disappearance of the
bright radical color in the presence of GSH, we calculated an apparent second-order
rate constant of about 105 M-1s-1. Addition
of a 100-fold excess of DMPD resulted in a decrease of the reaction rate
by a factor of 20. This experiment clearly indicated that the quinone diimine
is the reactive intermediate which reacts at pH 7.4 and 37°C with a
second-order rate constant of about 5 x 105 M-1s-1.
The main products formed from DMPD*+ and GSH were two isomeric
thioethers of DMPD, 2- and 3-(glutathione-S-yl)-N,N-dimethyl-p-phenylenediamine,
indicating a Michael addition reaction of GSH to the quinoid structure.
GSSG formation was negligible. Although the disproportionation reaction
of DMPD*+ is very fast, k = 2 x 106 M-1sec-1,
some saturation kinetics was observed when GSH was at high excess. A completely
different behaviour was found with the N,N,N´,N´-tetramethyl-p-phenylenediamine
radical cation (TMPD*+). The reaction velocity with GSH was independent
of the GSH concentration below 1 mM. Only at higher concentrations the expected
second-order reaction was found. Thioether formation at low concentrations
of GSH pointed to a Michael addition reaction to the quinoid compound. When
the GSH concentration was increased, the recovery of the thioether decreased.
Instead, GSSG formation increased.
Figure 2 compares the different behavior of the two Wurster's radicals.
The disparity is a result of the different rates of the disproportionation
of the radicals. With TMPD*+, the reaction is second-order at
low GSH concentrations and yields virtually constant amounts of the thioethers.
Only at high GSH concentrations does the disproportionation reaction become
rate limiting. This reaction is rather slow with TMPD*+ and is
obviously rate limiting during thioether formation. At increasing GSH, a
direct reaction of GSH with the radical is observed. During this process,
GSSG is formed and less quinone diimine is available for addition reactions.
Conceivably, at very high GSH concentrations, DMPD*+ will also
react directly with GSH at the expense of the thioethers. Hence, both phenylenediamine
radicals show two extremes. Figure 3 schematically compiles the whole continuum.
The left part is typical for DMPD*+, the right for TMPD*+.
Figure 2. Dependence of the velocity of radical disappearance
and thioether yields on GSH concentration during the interaction of DMPD*+
and TMPD*+, respectively, with GSH. Left panel: DMPD*+;
right panel: TMPD*+ DMPD*+, 2 µM; TMPD*+,
5 µM, pH 7.4, 37°C; argon. Adapted from Störle and Eyer (8,9,100).
Figure 3. Schematic
presentation of the dependence of the different reaction pathways of phenylenediamine
radical cations on the GSH concentration.
The tendency of the radicals to react directly with GSH was very weak.
With TMPD*+ a rate of 5 M-1sec-1 was determined
(9), whereas with DMPD*+, the rate is well below 102
M-1sec-1(8).
The mechanism of GSSG formation is still obscure. It may result from
the reaction of GSH with both the radical and the quinone diimine. In the
latter case, ipso adduct formation is conceivable, as already discussed
for other quinone imines (10-12). The situation in the former case
is more puzzling.
For thermodynamic reasons, TMPD*+ with a redox potential of
+ 0.27 V (13) is hardly able to be reduced by GSH with thiyl
radical formation (redox potential at pH 7.4 of about +0.9 V (14).
Hence, the equilibrium for GS* formation is about 4 x 10-10.
TMPD*+ + GS- <--------> TMPD
+ GS* [2]
(On the assumption of an effective pKa of 8.6 (15) for the thiol
dissociation, but ignoring other possible prototropic equilibria). Of course,
reactions that remove GS* from the equilibrium will drive the
reaction to the right. The most important reactions are the conjugation
of GS* with oxygen and GS-. The resulting radical
conjugates will be quickly eliminated by further reactions:
GS* + O2 --------> GSOO* [3]
GS* + GS- <--------> GSSG*-
[4]
GSSG*- + O2 --------> GSSG + O2*-
[5]
GSSG*- + TMPD*+ --------> GSSG
+ TMPD [6]
Because GSSG*- has a very low redox potential (-1.6 V),
the overall reaction can in fact result in slow TMPD*+ reduction
and GSSG formation. [For a more in-depth discussion of thiyl radical chemistry,
refer to Wardman (14)]. The possible involvement of the highly reactive
thiyl radicals may cause additional oxidative stress, as already reported
for p-phenetidine (16-18).
In conclusion, our experiments with N-alkylated p-phenylenediamines
indicate that the main reaction with thiols is brought about by the quinone
diimines that are usually in a rapid disproportionation equilibrium with
the radicals.
Aminophenoxyl Radicals
Our first contact with aminophenoxyl radicals dates back to the late
seventies, when we analyzed reactions of p-dimethylaminophenol (DMAP)
with oxyhemoglobin in the presence of GSH to imitate the red cell milieu.
Oxyhemoglobin and DMAP are cooxidized quickly with formation of a transient
N,N-dimethylamino-p-phenoxyl radical (19). This radical
also was generated by pulse radiolysis and--more conveniently--by reaction
with stoichiometric ferricyanide. The DMAP radical is quite stable, although
much less than the corresponding DMPD radical. It decays under physiologic
conditions with an apparent half-life of about 1 sec with formation of dimethylamine
and p-benzoquinone, indicating that the decay proceeds via the quinone
imine, which is in a rapid disproportionation equilibrium. Since the radical
of DMAP was nearly completely bleached by half an equivalent of GSH, we
assume that GSH reacts primarily with the quinone imine. In this reaction,
GSSG is hardly formed. Instead, thioethers are produced as expected from
Michael addition reactions to the quinone imine. Hence, the radical of DMAP
appears to react very similarly to the corresponding DMPD*+.
In contrast, the phenoxyl radical of paracetamol (NAPAP) (20) produced
in the horseradish peroxidase or prostaglandin H synthetase reaction, was
shown to react with GSH under formation of a thiyl radical as detected by
spin-trapping (17,18).
Quinone Imines
The reactions of GSH with the quinone imines from NAPAP, p-aminophenol,
and DMAP have been studied in some detail. With all three compounds, thioethers
were produced, but significant amounts of GSSG were found only with NAPAP
(21). The mechanism of GSSG formation has been attributed to intermediate
ipso adducts which, after attack of a second GSH molecule, yield GSSG and
NAPAP (10-12). During the reaction of the quinone imine of p-aminophenol
with GSH, sequential oxidation/addition reactions occur as revealed by a
variety of poly-substituted thioadducts. GSSG formation was not detected
(21). When the quinone imine of DMAP, prepared chemically by oxidation
with PbO2, reacted with GSH, some GSSG was found besides polysubstituted
thioadducts. This process also was examined in experiments where the oxidation
was brought about by oxyhemoglobin. As shown in Figure 4, DMAP disappeared
rapidly from incubates of oxyhemoglobin and glutathione with formation of
an intermediate mono-substituted adduct, followed by a bis- and tris-substituted
conjugate. Interestingly, GSSG formation proceeded only after a lag phase.
With the isolated mono- and bis-glutathione S-conjugates, we found
that significant GSSG formation occurred only during the reaction of 2,6-bis(glutathione-S-yl)4-dimethylaminophenol
with oxyhemoglobin and glutathione. Similar substituent effects on reduction
versus arylation have been reported for the quinone imines of dimethyl ring-substituted
NAPAP: the quinone imine disubstituted ortho to the keto function
gave only GSSG, whereas the compound disubstituted meta to the keto
function gave only the glutathione adduct (22). These data indicate
that substituents that influence the electron density at the quinoid system
will govern differently the rates of arylation and reduction (23).
Whether thiyl radicals are involved also, leading ultimately to GSSG, remains
to be established.
Figure 4. Formation
of glutathione S-conjugates during the oxyhemoglobin-mediated sequential
oxidation/addition reactions of 4-dimethylaminophenol (DMAP). Hemoglobin
(0.6 mM), with its sulfhydryl groups blocked by N-ethylmaleimide,
oxidized DMAP (0.1 mM) in the presence of GSH (0.5 mM) in 0.2 M sodium phosphate,
pH 7.4, at 37°C under air. The mono-, bis- and tris(glutathione-S-yl)
derivatives were determined by HPLC (101).
Toxicologic Impacts of the Reactions of Thiols with Quinone Imines
Alteration will occur in the glutathione status of those cells that are
involved in the activation of aromatic amines leading to arylaminyl and
aminophenoxyl radicals, and ultimately to quinoid compounds. Moderate GSSG
formation usually is well tolerated by cells with intact glutathione reductase
and glucose-6-phosphate dehydrogenase activities. Patients with glucose-6-phosphate
dehydrogenase deficiency and with the rare glutathione reductase deficiency,
however, are remarkably more sensitive. A typical example is the hematologic
disorder (i.e., hemolysis) resulting from the vicia faba aglykons
divicine and isouramil. As revealed by Winterbourn and coworkers (Figure
5) (24), the highly autoxidizable hydroxypyrimidines form radical
intermediates and quinoid derivatives in an autocatalytic manner, because
the dispropor- tionation reaction amplifies the autoxidizable radicals.
It is the particular interaction of GSH that prevents the exponential formation
of the radicals. Since the fully substituted hydroxypyrimidines do not allow
Michael addition reactions and therefore are not eliminated, GSH acts only
as a reductant.
Figure 5. Mechanisms
of oxidative stress induced by hydroxypyrimidine aglykons underlying favism.
Adapted from Winterbourn (24).
Aminophenols, like the more intensively investigated 4-dimethylaminophenol,
exert their toxic actions mainly as a result of arylation of cellular sulfhydryls.
The particular nephrotoxicity of DMAP is mainly due to depletion of essential
thiols within the mitochondria. DMAP depletes GSH and, more seriously, coenzyme
A levels with formation of tri-substituted mixed thioethers composed of
coenzyme A and GSH residues (25). Besides, enzymes with SH groups
essential for catalytic activity are inhibited (26 ). A particularly
illustrative example of the consequences of covalent binding of a quinoid
compound to protein sulfhydryls was detected in hemoglobin (Figure 6) (27).
DMAP, after activation by oxyhemoglobin, binds to the reactive SH groups
in cysteine 93 of the ß-chains. The resulting thioether autoxidizes
to give the quinoid derivative, which partly loses the dimethylamino group
on hydrolysis. In the last step, an imidazol nitrogen of the C-terminal
histidine residue binds to the quinoid thioether. By this process the protein
is cross-linked. The Perutz mechanism of conformation changes upon ligation
is blocked, and hemoglobin has lost its most important physiologic function:
it no longer liberates oxygen to the tissue, because of a 10-fold increased
oxygen affinity.
Figure 6. Scheme
of cross-link formation of the ß-chains of human hemoglobin by 4-dimethylaminophenol
(27).
What happens with the different thioethers that are produced with glutathione?
Glutathione S-conjugates produced in the liver are excreted predominantly
in the bile. Thus, the NAPAP thioether has been isolated and characterized
from rat bile (28). Glutathione S-conjugates from DMAP are
formed primarily in erythrocytes (29), actively exported through
the red cell membrane (30), and excreted in urine as premercapturic
acid (31,32). Similarly, p-aminophenol is activated in red
cells by oxyhemoglobin with formation of various glutathione S-conjugates,
which are extruded into the plasma (33). A variety of the glutathione
S-conjugates of 4-aminophenol and DMAP are still active in ferrihemoglobin
formation, produce reactive oxygen species, and are able to bind covalently
to hemoglobin (29,34,35). Recently, attention has been focused on
the particular target toxicity of glutathione S-conjugates in the
kidney. When glutathione synthesis was inhibited, 4-aminophenol was less
nephrotoxic to rats, and cannulation of the bile duct protected against
nephrotoxicity (36 ). Circumstantial evidence has been presented
that 4-amino-2-(glutathione-S-yl)phenol is excreted in the bile after
administration of 4-aminophenol to rats (37). This thioether autoxidizes
rapidly (21,34,38) and produces more ferrihemoglobin than the parent
4-aminophenol (35). Recently, it has been shown that another isomeric
thioether, 4-amino-3-(glutathione-S-yl)phenol is markedly more nephrotoxic
in Fischer 344 rats than 4-aminophenol (39). The kidney-specific
toxicity of autoxidizable glutathione S-conjugates results from the
capability of the proximal tubules to accumulate and process these conjugates
(37,40). Conceivably, oxidative stress by redox cycling and arylation
of cellular sulfhydryls (23,41) by the concentrated quinoid compounds
are the tubular cells' doom. It remains to be established whether glutathione
S-conjugates of aminophenols and phenylenediamines are also genotoxic
and contribute to renal carcinogenicity (40).
Mechanisms of the Reactions of Nitrosoarenes with Thiols
The reactions of thiols with nitrosoarenes are complex, and product formation
is dependent on thiol concentration, pH, and substituent effects (42-45).
With the model thiol, 1-thioglycerol (46,47), we could confirm the
reversible formation of a semimercaptallike intermediate, the existence
of which had been supposed already from kinetic measurements (43).
Meanwhile, the semimercaptal of 4-nitrosonitrobenzene and GSH also has been
isolated and characterized by NMR spectroscopy (48). The transient
semimercaptal can react in several ways (Figure 7):
Figure 7. Reaction
pathways of nitrosoarenes and glutathione.
a) The intermediate loses the hydroxyl group with formation of
an N-(thiol-S-yl)arylamine cation that may be trapped by reaction
of a solvent water molecule at the sulfur atom, giving rise to a sulfinamide
structure. This sequence has been confirmed with 18O-tracer experiments
(47,49).
b) A second thiolate anion reacts at the sulfur of the intermediate
semimercaptal, displacing the phenylhydroxylamine anion with formation of
GSSG. This reaction has been postulated repeatedly in agreement with reaction
kinetics and product pattern (43,44,46,48,49). Until now, however,
we have had no definite proof for this proposed reaction mechanism (47).
It should be noted that formation of phenylhydronitroxide radicals was
detected when nitrosobenzene was reacted with GSH, and it has been hypothesized
that nitrosobenzene is reduced to the phenylhydronitroxide radical with
formation of the thiyl radical (2,50). In fact, thiyl radical formation
was detected when low concentrations of nitrosobenzene (0.1 mM) were mixed
with 30 mM GSH in the presence of 100 mM DMPO as spin trap (K Maples, personal
communication). These results, however, cannot rule out definitively that
the phenylhydronitroxide radicals are formed by comproportionation of nitrosobenzene
and phenylhydroxylamine (51) and that thiyl radical formation occurs
during the reaction of the phenylhydronitroxide radical with GSH.
c) Formation of a sulfenamide, N-(thiol-S-yl)-arylamine,
was detected with 2-nitrosofluorene and GSH (52), with nitrosobenzene
and 1-thioglycerol (46 ), and with 4-nitrosophenetol and GSH (53,
54). Sulfenamide formation was proposed initially to occur through a
mercaptal intermediate, ArN(SR)2, followed by thiolytic cleavage
(44). Such a reaction became less probable later on, because reaction
of ArN(OH)SR with different thiols R´SH did not yield ArNHSR´
but always ArNHSR (46 ). Kazanis et al. (49) in their brilliant
work offer an attractive explanation for sulfenamide formation from the
N-(thiol-S-yl)-arylamine cation: nucleophilic addition to
the ring, particularly when ¼-donors are in para position, can
lead to a thioadduct incapable of aromatization. This adduct can be cleaved
thiolytically and forms the sulfenamide. The latter can be cleaved thiolytically
once more with formation of the arylamine. In the resonance-stabilized N-(thiol-S-yl)-arylamines
with strong ¼-donors, the positive charge is highly delocalized to
the ring carbon atoms thereby facilitating other addition reactions. We
have isolated such products from 4-nitrosophenetol and GSH, 4-ethoxy-2-(glutathione-S-yl)-aniline
and a variety of polycyclic compounds (47,54,55).
Investigation of substituent effects have indicated clearly a Hammett
correlation of the reactivity and the product pattern of p-substituted
nitrosoarenes in their reactions with GSH (45,49,55). Thus, 4-nitrosonitrobenzene
gives only the phenylhydroxylamine via the semimercaptal (48). The
same holds true for another nitrosoarene with strong electron-withdrawing
groups--4-nitrosoacetophenone (45). The intermediary nitrosobenzene
gives phenylhydroxyl- amine, the sulfinamide, and the sulfenamide. On the
other end of the Hammett scale, 4-nitrosophenetol and 4-nitroso-N,N-dimethylaniline
with their strong pi-donating substituents yield no phenyl- hydroxylamine,
but ultimately yield the amine besides the other adducts mentioned above
(45,47,54,55).
Toxicologic Impacts of the Reactions of Thiols with
Nitroso Compounds
Nitrosoimidazoles
Nitroheterocyclic compounds like the nitroimidazoles are used worldwide
as antimicrobial, antiparasitic, and radiosensitizing agents. Metabolic
activation, mainly by nitro group reduction, seems to be a prerequisite
for the biologic effects (56 ). The influence of thiols, mainly GSH,
on the toxicity, mutagenicity, and DNA damage has revealed complex mechanisms.
The 2-hydroxylaminoimidazoles apparently react directly with thiols to form
unstable 2-N-(glutathione-S-yl) conjugates which in the presence
of excess GSH produce two stable 4-C- and 5-C-(glutathione-S-yl)
conjugates, possibly with a nitrenium ion as intermediate (57-60).
These data indicate that thiols may both activate and inactivate penultimate
reactive nitroimidazole compounds.
GSH and cysteine enhanced the incorporation of labeled 1-methyl-4-phenyl-5nitrosoimidazole
into 
-phage DNA and polynucleotides (61). The authors
concluded that a semimercaptal-like intermediate might yield a highly reactive
cation that binds to DNA. These data suggest that the nitrosoimidazole might
still be a penultimate reactive intermediate in the bioactivation of nitroimidazoles,
which by interaction with thiols gives the ultimate reactive species that
binds to DNA.
1-Methyl-2-nitrosoimidazole was two orders of magnitude more toxic to
CHO cells than the nitro and hydroxylamine compound. Circumstantial evidence
suggested that GSH may reduce cytotoxicity (62). Similar observations
were reported with HT-29 human colon cancer cells. Depletion of cellular
GSH with buthionine sulfoximine before incubation with the nitrosoimidazole
resulted in enhanced susceptibility (63). In chemical systems, 1methyl-2-nitrosoimidazole
reacted rapidly with GSH, with formation of GSSG, the hydroxylamine, and
imidazole-GSH adducts. Mixtures of the nitrosoimidazole with GSH gave rise
to DNA strand breaks in the plasmid assay (64). Since the DNA damage
by the nitrosoimidazole in HT-29 colon cancer cells probably was not a result
of a direct interaction of the nitroso compound, a possible activating effect
of GSH to yield the ultimate electrophile (which is not the hydroxylamine)
was discussed (65). All these hypotheses have to be confirmed, but
it appears that GSH might have an ambiguous role in the reactions with nitrosoimidazoles.
Probably, steady state concentrations of both the metastable intermediates
and GSH at sensitive targets critically control activating and inactivating
reactions.
Heterocyclic Nitroso Compounds from Protein Pyrolysates
Sugimura and co-workers have found numerous mutagenic compounds in broiled
fish and meat, and in pyrolysates of protein and amino acids. These mutagens
are heterocyclic amines and exhibit mutagenicity in the presence of S9 mix
(66 ). All these mutagens, derivatives of pyridoindole (Trp-P-1 and
Trp-P-2), pyridoimidazole (Glu-P-1 and Glu-P-2), and imidazoquinoline (IQ)
are N-hydroxylated at an exocyclic amino group to form proximate
reactive compounds.
Thiols, particularly GSH, modify the mutagenic activity and covalent
binding to DNA by at least two mechanisms. The first one involves glutathione
S-transferase, which in the case of N-OH-Trp-P-2 produces a stable
C-conjugate and two labile N-conjugates, a sulfenamide and,
to the best of our knowledge, a hitherto unidentified derivative that easily
liberates the parent N-OH-Trp-P-2. This last compound was found to be outstandingly
mutagenic (67,68). The second pathway starts with the nitroso derivative
leading to the putative semimercaptal-like intermediate (69), a sulfenamide,
and a sulfinamide (68). Most interestingly, also a N-hydroxysulfonamide
from NO-Glu-P-1 was apparently formed (70), which may have derived
from reaction of the (liberated) glutathione sulfinic acid with NO-Glu-P-1.
Such a reaction has been reported to occur with nitrosoarenes and arenesulfinic
acids (71). A direct reaction of nitroso compounds of interest with
authentic glutathione sulÞnic acid under physiologic conditions, however,
remains to be established. No reaction between N-OH-Glu-P-1 and GSH
was detected (70). At present, it appears that the interaction of
GSH with the nitroso heterocyclic pyrolysates is mainly a detoxication reaction.
Nitrosoarenes
A chemical intermediate widely used in the synthesis of certain dyes
and explosives, m-dinitrobenzene has been found to cause testicular
toxicity, with the Sertoli cells being the initial target. The toxicity
is probably mediated through m-nitrosonitrobenzene. Depletion of
cellular GSH with diethylmaleate made the cultured rat Sertoli cells more
susceptible to damage by m-dinitrobenzene and m-nitrosonitrobenzene,
while cysteamine reduced the toxicity. These data suggest that GSH interferes
with metabolically formed reactive species, probably by scavenging the nitrosoarene
(48,72).
2-Nitrosofluorene is a potent direct-acting mutagen that is derived from
biotransformation of the well-known carcinogen N-acetyl-2-aminofluorene
(73). The mutagenicity of 2-nitrosoþuorene is known to be inhibited
by inclusion of GSH in the assay media. Interestingly, the reaction of 2-nitrosoþuorene
with GSH, which yields a water-soluble product that liberated 2-aminofluorene
under acidic conditions, is probably the first example of this class of
reactions reported in the literature (74). Later on, others have
elucidated the reactions more intensively (44, 52) and found hydroxylamine,
sulfinamide, and sulfenamide formation. In addition, slow formation of the
sulfenamide from the interaction of N-hydroxy-2-aminofluorene with
GSH was reported, and a nucleophilic displacement reaction was suggested
to be underlying (52). Because this reaction was also observed in
the presence of ascorbic acid, interference of 2-nitrosofluorene via autoxidation
or disproportionation was excluded. Conceivably, N-O cleavage is
facilitated by the stabilizing effect of the phenyl residue that may act
as a strong ¼-donor (49). The resulting nitrenium ion may then
react with GSH to form the sulfenamide. In fact, N-hydroxy-2-aminofluorene
is quite electrophilic and reacts readily with DNA and deoxyguanosine, even
at neutral pH (FF Kadlubar, personal communication, 1984). 4-Nitrosophenetol
bound directly to tRNA (75) but only weakly to calf thymus DNA, whereas
N-hydroxy-4-phenetidine bound readily (53). GSH was without
effect on binding of 4-nitrosophenetol.
Studying toxic effects of p-substituted nitrosoarenes with widely
varying Hammett constants that induce cellular damage in isolated rat hepatocytes,
O'Brien and co-workers detected dichotomic modulating effects of GSH (41).
GSH depletion by diethylmaleate decreased cytotoxicity by nitrosobenzene
and 4-chloronitrosobenzene but increased cytotoxicity with 4-nitrosoacetophenone
and 4-nitroso-N,N-dimethylaniline. From these data, it was concluded
that those nitrosoarenes capable of forming sulfinamides with GSH (45)
may produce particularly toxic intermediates (e.g., the sulfinamide itself)
(76 ) or, more likely, the electrophilic arylnitrenium cations as
proposed by McClelland's group (49). Hence, these observations point
to mechanisms whereby GSH enhances toxicity. The protective effect of GSH
on 4-nitrosoacetophenone toxicity might be a result of the very rapid reduction
rates with formation of the hydroxylamine and GSSG [k=105 M-1sec-1
(45)]. A similar mechanism in 4-nitroso-N,N-dimethylaniline
toxicity is less likely, since the reaction with GSH is sluggish [k=1.5
M-1sec-1 (45)]. In this particular case, another
detoxication mechanism appears to act: conjugation with GSH is brought about
by hepatic glutathione S-transferase (77).
A peculiar toxic action of nitrosoarenes was observed during liver perfusion.
4Nitrosophenetol (78) and 4-nitroso-N,N-dimethylaniline inhibited
glutathione and GSSG excretion into bile, while nitrosobenzene and 4-nitrosoacetophenone
were inactive (3). Such an inhibition has been observed also in erythrocytes,
where 4-nitrosophenetol inhibited the export of the model glutathione S-conjugate,
S-(2,4-dinitrophenyl)-glutathione, in a competitive manner. The competition
is probably caused by glutathione S-conjugates of 4-nitrosophenetol
(55). 4-Nitrosophenetol and the bicyclic derivative 4-ethoxy-4´-nitrosodiphenylamine
have been shown to be cytotoxic to isolated rat hepatocytes with GSH depletion
(79).
The most obvious effects of GSH on cytotoxicity of nitrosoarenes are
observed in the blood. The methemoglobinemia and hemolysis brought about
by nitrosoarenes in the Kiese cycle, where nitrosoarenes are enzymically
reduced and the phenylhydroxylamines are cooxidized to yield ferrihemoglobin
and the parent nitrosoarene (80-82), are markedly influenced by reactions
with thiols. GSH decreases the extent of ferrihemoglobin formation and increases
the clearance of the blood from the nitrosoarenes (3,83-86 ). Such
an example is shown in Figure 8, where GSH markedly diminished ferrihemoglobin
formation by eliminating 4-nitrosophenetol from the Kiese cycle.
Figure 8. Influence
of GSH on the decrease in 4-nitrosophenetol (NOPt), ferrihemoglobin (HbFe3+),
and phenetidine (NH2Pt) formation in the reaction of nitrosophenetol
with hemolysate, fortified by a NADPH-regenerating system (3).
The importance of the clearing function of blood to protect other sensitive
target organs has been postulated earlier (84) and confirmed in the
case of nitrosochloramphenicol. This bacterial metabolite of chloramphenicol
has been accused to produce the occasionally observed stem cell damage (87).
However, nitrosochloramphenicol reacts very rapidly with GSH in chemical
systems (88) and in the blood (85), so that the reactive compound
was calculated to be degraded within a few seconds. Hence, nitrosochloramphenicol,
formed by microorganisms in the intestine or produced in the liver, will
be degraded in blood before it can reach the bone marrow (85,89).
Interestingly, another bacterial chloramphenicol metabolite emerged as a
favorite proximate toxic candidate, dehydrochloramphenicol, a compound with
a propiophenone moiety (89) (Figure 9). This compound is fairly stable
in the blood and can reach the bone marrow cells. Dehydrochloramphenicol
itself inhibits myeloid colony growth. Perhaps the most important aspect
of dehydrochloramphenicol is that, in contrast to chloramphenicol, it is
readily reduced by human bone marrow homogenate, even under aerobic conditions
(90). One can assume that the 4nitrosopropiophenone will react with
GSH similarly to 4-nitrosoacetophenone to give the hydroxylamine and GSSG,
thereby inducing a marked oxidative stress. Further investigations will
show whether this metabolic pathway is indeed responsible for the chloramphenicol
induced aplastic anemia.
Figure 9. Structural
formulae of chloramphenicol and putative myelotoxic metabolites.
A variety of arylamine drugs induce idiosyncratic reactions like lupus
erythematodes and agranulocytosis. Procainamide is associated with a relatively
high incidence of both idiosyncrasies. It has been suggested that myeloperoxidase-mediated
reactions might be involved. The hydroxylamine and nitroso derivatives of
procainamide were produced by activated human neutrophiles and in the presence
of myeloperoxidase/ hydrogen peroxide. The nitroso compound was found to
react rapidly with thiols to give a sulfinamide (91). In addition,
covalent binding to proteins was observed. Such a reaction sequence, which
produces a hapten-macromolecule adduct, will ultimately induce antibody
formation and give rise to an immune system-mediated idiosyncratic drug
reaction. Similar reactions may be underlying other idiosyncrasies observed
with dapsone, sulfonamides, metoclopramide, and p-aminosalicyclic
acid (92,93).
Finally, and notably, nitrosoarene reactions with SH groups in human
hemoglobin are particularly worthy to mention because of the genius loci.
Neumann and coworkers have undertaken many efforts during the last decade
to use hemoglobin adducts as a dose monitor in exposure control of aromatic
amines and their relationship to risk assessment (44,94-96 ). The
method makes use of sulfinamide formation of nitrosoarenes with the exposed
SH group of cysteine ß93 in human hemoglobin. These adducts are rather
stable to provide an integral dose monitor for the internal exposure to
and the extent of metabolic activation of aromatic amines. The phenylhydroxylamines
are cooxidized by oxyhemoglobin, and a fraction of the nitroso compound
can react with the hemoglobin sulfhydryls. The precipitated hemoglobin of
washed red cells is hydrolyzed, and the liberated amine is determined. This
method may also allow the identification of persons with high individual
risk (acetylator status, oxidative phenotype, G6PDH deficiency).
Of course, direct comparison of the hemoglobin binding indices of various
aromatic amines (or nitroarenes) has to consider the widely varying bioavailability
of the nitrosoarenes for binding, because steric and substituent effects
markedly influence the yield of sulfinamide formation with hemoglobin SH
groups (86,97), topics that have already been discussed.
Conclusion
The study of the reactions of oxidatively activated arylamines with thiols
has revealed a puzzling variety of reaction types. Some pathways lead to
rather stable products that are excreted with urine or bile and can be regarded
as water-soluble, inactive conjugates. Other pathways lead to metastable
derivatives that can be reactivated in particular target cells. Autoxidizable
thioethers are concentrated in the kidney; labile thiol adducts of nitroso
compounds may liberate carbonium/nitrenium ions that exhibit genotoxic activity.
Covalent binding to proteins can inactivate vital enzymes and may lead to
haptenization, followed by an immune response. Last, covalently bound adducts
to proteins can pose a latent risk when the adducts are degraded, either
in the body or in processed food from animals that have been treated with
antibiotics (e.g., chloramphenicol, nitrofurans, or nitroimidazoles) or
from agricultural products contaminated with herbicides, insecticides, etc.
The rather new public interest in the toxicity of (drug) residues bound
covalently to cysteine (98,99) will probably help toxicologists remain
employed well into the future.
REFERENCES
1. Kriek E, Westra JG, Welling M. Reaction of N-oxidation
products of aromatic amines with nucleic acids. In: Biological Oxidation
of Nitrogen in Organic Molecules: Chemistry, Toxicology and Pharmacology
(Gorrod JW, Damani LA, eds). Chichester, England:Horwood E, 1985;367-376.
2. Maples KR, Eyer P, Mason RP. Aniline-, phenylhydroxylamine-,
nitrosobenzene-, and nitrobenzene-induced hemoglobin thiyl free radical
formation in vivo and in vitro. Mol Pharmacol 37:311-318 (1990).
3. Eyer P. Reactions of nitrosoarenes with sulphydryl groups:
reaction mechanism and biological significance. In: Biological Oxidation
of Nitrogen in Organic Molecules: Chemistry, Toxicology and Pharmacology
(Gorrod JW, Damani LA, eds). Chichester, England:Horwood E, 1985;386-399.
4. Mason RP. Free-radical intermediates in the metabolism
of toxic chemicals. In: Free Radicals in Biology, Vol V ( Pryor WA, ed).
Orlando, Florida:Academic Press, 1982;161-222.
5. Eyer P. Activation of aromatic amines by oxyhemoglobin.
In: N-Oxidation of Drugs, Biochemistry, Pharmacology and Toxicology
(Hlavica P, Damani LA, eds). London:Chapman & Hall, 1991;371-391.
6. Zenser TV, Cohen SM, Mattammal MB, Wise RW, Rapp NS,
Davis BB. Prostaglandin hydroperoxidase-catalyzed activation of certain
N-substituted aryl renal and bladder carcinogens. Environ Health
Perspect 49:33-41 (1983).
7. Michaelis L, Schubert MP, Granick S. The free radicals
of the type of Wurster's salts. J Am Chem Soc 61:1981-1992 (1939).
8. Störle C, Eyer P. Reactions of the Wurster's red
radical cation with hemoglobin and glutathione during the cooxidation of
N,N-dimethyl-p-phenylenediamine and oxyhemoglobin in human
red cells. Chem Biol Interact 83:271-291 (1992).
9. Störle C, Eyer P. Reactions of the Wurster's blue
radical cation with thiols, and some properties of the reaction products.
Chem Biol Interact 78:333-346 (1991).
10. Fernando CR, Calder IC, Ham KN. Studies on the mechanism
of toxicity of acetaminophen. Synthesis and reactions of N-acetyl-2,6-dimethyl-
and N-acetyl-3,5-dimethyl-p-benzoquinone imines. J Med Chem
23:1153-1158 (1980).
11. Rundgren M, Porubek DJ, Harvison PJ, Cotgreave IA,
Moldéus P, Nelson SD. Comparative cytotoxic effects of N-acetyl-p-benzoquinone
imine and two dimethylated analogues. Mol Pharmacol 34:566-572 (1988).
12. Axworthy CA, Hoffmann K-J, Streeter AJ, Calleman CJ,
Pascoe GA, Baillie TA. Covalent binding of acetaminophen to mouse hemoglobin.
Identification of major and minor adducts formed in vivo and implications
for the nature of the arylating metabolites. Chem Biol Interact 68:99-116
(1988).
13. Steenken S, Neta P. One-electron redox potentials of
phenols. Hydroxy- and aminophenols, and related compounds of biological
interest. J Phys Chem 86:3661-3667 (1982).
14. Wardman P. Conjugation and oxidation of glutathione
via thiyl free radicals. In: Glutathione Conjugation (Sies H, Ketterer B,
eds). London:Academic Press, 1988;43-72.
15. Eyer P, Podhradsky D. Evaluation of the micromethod
for determination of glutathione using enzymatic cycling and Ellman's reagent.
Anal Biochem 153:57-66 (1986).
16. Larsson R, Ross D, Nordenskjöld M, Lindeke B,
Olsson LI, Moldéus P. Reactive products formed by peroxidase catalyzed
oxidation of p-phenetidine. Chem Biol Interact 52:1-14 (1984).
17. Ross D, Albano E, Nilsson U, Moldéus P. Thiyl
radicals--formation during peroxidase-catalyzed metabolism of acetaminophen
in the presence of thiols. Biochem Biophys Res Comm 125:109-115 (1984).
18. Ross D, Larsson R, Andersson B, Nilsson U, Lindqvist
T, Lindeke B, Moldéus P. The oxidation of p-phenetidine by
horseradish peroxidase and prostaglandin synthase and the fate of glutathione
during such oxidations. Biochem Pharmacol 34:343-351 (1985).
19. Eyer P, Lengfelder E. Radical formation during autoxidation
of 4-dimethylaminophenol and some properties of the reaction products. Biochem
Pharmacol 33:1005-1013 (1984).
20. West PR, Harman LS, Josephy PD, Mason RP. Acetaminophen:
enzymatic formation of a transient phenoxyl free radical. Biochem Pharmacol
33:2933-2936 (1984).
21. Eckert K-G, Eyer P, Sonnenbichler J, Zetl I. Activation
and detoxication of aminophenols. II. Synthesis and structural elucidation
of various thiol addition products of 1,4-benzoquinone imine and N-acetyl-1,4-benzoquinone
imine. Xenobiotica 20:333-350 (1990).
22. Rosen GM, Rauckman EJ, Ellington SP, Dahlin DC, Christie
JL, Nelson SD. Reduction and glutathione conjugation reactions of N-acetyl-p-benzoquinoneimine
and two dimethylated analogues. Mol Pharmacol 25:151-157 (1984).
23. Monks TJ, Hanzlik RP, Cohen GM, Ross D, Graham DG.
Quinone chemistry and toxicity. Toxicol Appl Pharmacol 112:2-16 (1992).
24. Winterbourn CC. Inhibition of autoxidation of divicine
and isouramil by the combination of superoxide dismutase and reduced glutathione.
Arch Biochem Biophys 271:447-455 (1989).
25. Eckert K-G, Elbers FR, Eyer P. Depletion of mitochondrial
coenzyme A and glutathione by 4-dimethylaminophenol and formation of mixed
thioethers. Biochem Pharmacol 38:3253-3259 (1989).
26. Szinicz L, Weger N, Schneiderhahn W, Kiese M. Nephrotoxicity
of aminophenols: effects of 4-dimethylaminophenol on isolated rat kidney
tubules. Arch Toxicol 42:63-67 (1979).
27. Eyer P, Lierheimer E, Strosar M. Site and mechanism
of covalent binding of 4-dimethylaminophenol to human hemoglobin, and its
implications to the functional properties. Mol Pharmacol 24:282-290 (1983).
28. Hinson JA, Monks TJ, Hong M, Highet J, Pohl LR. 3(Glutathion-S-yl)acetaminophen:
a biliary metabolite of acetaminophen. Drug Metab Dispos 10:47-50 (1982).
29. Eyer P, Kiese M. Biotransformation of 4-dimethylaminophenol:
reaction with glutathione, and some properties of the reaction products.
Chem Biol Interact 14:165-178 (1976).
30. Eckert K-G, Eyer P. Formation and transport of xenobiotic
glutathione S-conjugates in red cells. Biochem Pharmacol 35:325-329 (1986).
31. Eyer P, Gaber H. Biotransformation of 4-dimethylaminophenol
in the dog. Biochem Pharmacol 27:2215-2221 (1978).
32. Klimmek R, Krettek C, Szinicz L, Eyer P, Weger N. Effects
and biotransformation of 4-dimethylaminophenol in man and dog. Arch Toxicol
53:275-288 (1983).
33. Eckert K-G. The metabolism of aminophenols in erythrocytes.
Xenobiotica 18:1319-1326 (1988).
34. Eckert K-G, Eyer P. Thiol-mediated activation of hydroquinones
and aminophenols, the drawback of thioether formation. International Symposium
on Mechanisms of Tissue Specific Toxicity. Würzburg, Germany, 1989;10.
35. Eckert K-G, Eyer P. Formation of reactive oxygen species
during the thiol-mediated activation of 1,4-hydroquinone, aminophenols and
paracetamol. Naunyn Schmiedeberg's Arch Pharmacol 341(Suppl R15):1990.
36. Gartland KPR, Eason CT, Bonner FW, Nicholson JK. Effects
of biliary cannulation and buthionine sulphoximine pretreatment on the nephrotoxicity
of para-aminophenol in the Fischer 344 rat. Arch Toxicol 64:14-25 (1990).
37. Koob M, Dekant W. Bioactivation of xenobiotics by formation
of toxic glutathione conjugates. Chem Biol Interact 77:107-136 (1991).
38. Eckert K-G, Eyer, Sonnenbichler J, Zetl I. Thioether
formation-activation or detoxication? Arch Toxicol (Suppl)13:287-290 (1989).
39. Fowler LM, Moore RB, Foster JR, Lock EA. Nephrotoxicity
of 4-aminophenol glutathione conjugate. Hum Exp Toxicol 10:451-459 (1991).
40. Monks TJ, Anders MW, Dekant W, Stevens JL, Lau SS,
van Bladeren PJ. Glutathione conjugate mediated toxicities. Toxicol Appl
Pharmacol 106:1-19 (1990).
41. O'Brien PJ, Wong WC, Silva J, Khan S. Toxicity of nitrobenzene
compounds towards isolated hepatocytes: dependence on reduction potential.
Xenobiotica 20:945-955 (1990).
42. Neumann H-G, Metzler M, Töpner W. Metabolic activation
of diethylstilbestrol and aminostilbene-derivatives. Arch Toxicol 39:21-30
(1977).
43. Eyer P. Reactions of nitrosobenzene with reduced glutathione.
Chem Biol Interact 24:227-239 (1979).
44. Dölle B, Töpner W, Neumann H-G. Reaction
of arylnitroso compounds with mercaptans. Xenobiotica 10:527-536 (1980).
45. Diepold C, Eyer P, Kampffmeyer H, Reinhardt K. Reactions
of aromatic compounds with thiols. In: Biological Reactive Intermediates
II. Chemical Mechanisms and Biological Effects, Vol B (Snyder R, Parke DV,
Kocsis JJ, Jollow DJ, Gibson CG, Witmer CM, eds). New York:Plenum Press,
1982;1173-1181.
46. Klehr H, Eyer P, Schäfer W. On the mechanism of
reactions of nitrosoarenes with thiols. Formation of a common intermediate
"semimercaptal." Biol Chem Hoppe-Seyler 366:755-766 (1985).
47. Klehr H. Zum Mechanismus der Reaktionen von Nitrosoarenen
mit Thiolen. Thesis. München, Germany: Ludwig Maximilians Universität,
1988.
48. Ellis MK, Hill S, Matthew P, Foster D. Reactions of
nitrosonitrobenzenes with biological thiols: identification and reactivity
of glutathione S-yl conjugates. Chem Biol Interact 82:151-163 (1992).
49. Kazanis S, McClelland RA. Electrophilic intermediate
in the reaction of glutathione and nitrosoarenes. J Am Chem Soc 114:3052-3059
(1992).
50. Takahashi N, Fischer V, Schreiber J, Mason RP. An ESR
study of nonenzymatic reactions of nitroso compounds with biological reducing
agents. Free Radic Res Commun 4:351-358 (1988).
51. Lenk W, Riedl M. N-Hydroxy-N-arylacetamides.
V. Differences in the mechanism of haemoglobin oxidation in vitro
by N-hydroxy-4-chloroacetanilide and N-hydroxy-4-chloroaniline.
Xenobiotica 19:453-475 (1989).
52. Mulder GJ, Unruh LE, Evans FE, Ketterer B, Kadlubar
FF. Formation and identification of glutathione conjugates from 2-nitrosofluorene
and N-hydroxy-2-aminofluorene. Chem Biol Interact 39:111-127 (1982).
53. Mulder G, Kadlubar FF, Mays JB, Hinson JA. Reaction
of mutagenic phenacetin metabolites with glutathione and DNA. Possible implications
for toxicity. Mol Pharmacol 26:342-347 (1984).
54. Klehr H, Eyer P, Schäfer W. Formation of 4-ethoxy-4´-nitrosodiphenylamine
in the reaction of the phenacetin metabolite 4-nitrosophenetol with glutathione.
Biol Chem Hoppe-Seyler 368:895-902 (1987).
55. Gallemann D, Eyer P. Additional pathways of S-conjugate
formation during the interaction of thiols with nitrosoarenes bearing ¼-donating
substituents. Environ Health Perspect 101 (Suppl 10) (1993).
56. Kedderis GL, Miwa GT. The metabolic activation of nitroheterocyclic
therapeutic agents. Drug Metab Rev 19:33-62 (1988).
57. Varghese AJ. Glutathione conjugates of misonidazole.
Biochem Biophys Res Commun 112:1013-1020 (1983).
58. Varghese AJ, Whitmore GF. Properties of 2-hydroxylaminoimidazoles
and their implications for the biological effects of 2-nitroimidazoles.
Chem Biol Interact 56:269-287 (1985).
59. Whitmore GF, Varghese AJ. The biological properties
of reduced nitroheterocyclics and possible underlying biochemical mechanisms.
Biochem Pharmacol 35:97-103 (1986).
60. Bolton JL, McClelland RA. Kinetics and mechanism of
the decomposition in aqueous solutions of 2-(hydroxyamino)imidazoles. J
Am Chem Soc 111:8172-8181 (1989).
61. Ehlhardt WJ, Goldman P. Thiol-mediated incorporation
of radiolabel from 1-[14C]-methyl-4-phenyl-5-nitrosoimidazole
into DNA. Biochem Pharmacol 38:1175-1180 (1989).
62. Noss MB, Panicucci R, McClelland RA, Rauth AM. Preparation,
toxicity and mutagenicity of 1-methyl-2-nitrosoimidazole. Biochem Pharmacol
37:2585-2593 (1988).
63. Mulcahy RT, Gipp JJ, Ublacker GA, Panicucci R, McClelland
RA. Cytotoxicity and glutathione depletion by 1-methyl-2-nitrosoimidazole
in human colon cancer cells. Biochem Pharmacol 38:1667-1671 (1989).
64. Berube LR, Farah S, McClelland RA, Rauth AM. Effect
of 1methyl-2-nitrosoimidazole on intracellular thiols and calcium levels
in Chinese hamster ovary cells. Biochem Pharmacol 42:2153-2161 (1991).
65. Gipp JJ, McClelland RA, Mulcahy RT. DNA damage induced
in HT-29 colon cancer cells by exposure to 1-methyl-2-nitrosoimidazole,
a reductive metabolite of 1-methyl-2-nitroimidazole. Biochem Pharmacol Suppl
42:127-133 (1991).
66. Sugimura T, Sato S, Wakabayashi K. Mutagens/carcinogens
in pyrolysates of amino acids and proteins and cooked foods: heterocyclic
aromatic amines. In: Chemical Induction of Cancer, Vol IIIC (Lai Y-T, Arcos
JC, Argus MF, eds). San Diego, CA:Academic Press, 1988;681-710.
67. Saito K, Yamazoe Y, Kamataki T, Kato R. Activation
and detoxication of N-hydroxy-Trp-P-2 by glutathione and glutathione
transferases. Carcinogenesis 4:1551-1557 (1983).
68. Kato R. Metabolic activation of mutagenic heterocyclic
aromatic amines from protein pyrolysates. Crit Rev Toxicol 16:307-348 (1986).
69. Saito K, Kato R. Glutathione conjugation of arylnitroso
compound: detection and monitoring labile intermediates in situ inside a
fast atom bombardment mass spectrometer. Biochem Biophys Res Commun 124:1-5
(1984).
70. Umemoto A, Grivas S, Yamaizumi Z, Sato S, Sugimura
T. Nonenzymatic glutathione conjugation of 2-nitroso-6-methyldipyrido [1,2-a:
3´,2´-d] imidazole (NO-Glu-P-1) in vitro: N-hydroxy-sulfonamide,
a new binding form of arylnitroso compounds and thiols. Chem Biol Interact
68:57-69 (1988).
71. Darchen S, Moinet C. Arenesulphinic acids. Nitroso
protecting reagents applicable to some nitrosoarenes. J Chem Soc Chem Commun
820 (1976).
72. Cave DA, Foster PMD. Modulation of m-dinitrobenzene
and m-nitrosonitrobenzene toxicity in rat Sertoli-germ cell cocultures.
Fundam Appl Toxicol 14:199-207 (1990).
73. Ames BN, Gurney EG, Miller JA, Bartsch H. Carcinogens
as frameshift mutagens: metabolites and derivatives of 2-acetylaminofluorene
and other aromatic amine carcinogens. Proc Natl Acad Sci USA 69:3128-3232
(1972).
74. Lotlikar PD, Miller EC, Miller JA, Margreth A. The
enzymatic reduction of the N-hydroxy derivatives of 2-acetylaminofluorene
and related carcinogens by tissue preparations. Cancer Res 25:1743-1752
(1965).
75. Vaught JB, McGarvey PM, Lee M, Garner CD, Wang CY,
Linsmaier-Bednar EM, King CM. Activation of N-hydroxyphenacetin to
mutagenic and nucleic acid-binding metabolites by acyltransfer, deacylation
and sulfate conjugation. Cancer Res 41:3424-3429 (1981).
76. Silva JM, Jatoe SD, O'Brien PJ. Glutathione may mediate
the cytotoxicity of some nitrosoarenes. Prog Pharmacol Clin Pharmacol 8:289-299
(1991).
77. Silva JM, O'Brien PJ. Detoxification of 4-nitroso-N,N-dimethylaniline
in isolated rat hepatocytes. Prog Pharmacol Clin Pharmacol 8:301-308 (1991).
78. Eyer P, Kampffmeyer H. Inhibition of glutathione excretion,
bile flow, and alterations of the glutathione status by 4-nitrosophenetol
during liver perfusion. Chem Biol Interact 42:209-223 (1982).
79. Lindqvist T, Moldéus P, Lindeke B. Cellular
effects of some metabolic oxidation products pertinent to 4-ethoxyaniline.
Pharmacol Toxicol 69:117-121 (1991).
80. Kiese M, Reinwein D, Waller HD. Kinetik der Hämiglobinbildung.
IV. Die Hämiglobinbildung durch Phenylhydroxylamin und Nitrosobenzol
in roten Zellen in vitro. Arch Exp Path Pharmakol 210:393-398 (1950).
81. Kiese M. Methemoglobinemia: A Comprehensive Treatise.
Cleveland, Ohio:CRC Press, 1974.
82. McMillan DC, Bradshaw TP, Hinson JA, Jollow DJ. Role
of metabolites in propanil-induced hemolytic anemia. Toxicol Appl Pharmacol
110:70-78 (1991).
83. Eyer P, Lierheimer E. Biotransformation of nitrosobenzene
in the red cell and the role of glutathione. Xenobiotica 10:517-526 (1980).
84. Eyer P. The red cell as a sensitive target for activated
toxic arylamines. Arch Toxicol (Suppl) 6:3-12 (1983).
85. Eyer P, Lierheimer E, Schneller M. Reactions of nitrosochlor-amphenicol
in blood. Biochem Pharmacol 33:2299-2308 (1984).
86. Eyer P. Detoxication of N-oxygenated arylamines
in erythrocytes. An overview. Xenobiotica 18:1327-1333 (1988).
87. Yunis AA, Miller AM, Salem Z, Arimura GK. Chloramphenicol
toxicity: pathogenetic mechanisms and the role of the p-NO2
in aplastic anemia. Clin Toxicol 17:359-373 (1980).
88. Eyer P, Schneller M. Reactions of the nitroso analogue
of chloramphenicol with reduced glutathione. Biochem Pharmacol 32:1029-1036
(1983).
89. Abou-Khalil WH, Yunis AA, Abou-Khalil S. Stability
of chloramphenicol metabolites in human blood and liver as determined by
high-performance liquid chromatography. Pharmacology 36:272-278 (1988).
90. Yunis AA. Chloramphenicol: relation of structure to
activity and toxicity. Annu Rev Pharmacol Toxicol 28:83-100 (1988).
91. Uetrecht JP. Reactivity and possible significance of
hydroxylamine and nitroso metabolites of procainamide. J Pharmacol Exp Ther
232: 420-425 (1984).
92. Cribb AE, Miller M, Leeder JS, Hill J, Spielberg SP.
Reactions of the nitroso and hydroxylamine metabolites of sulfamethoxazole
with reduced glutathione. Drug Metab Dispos 19:900-906 (1991).
93. Uetrecht JP. The role of leukocyte-generated reactive
metabolites in the pathogenesis of idiosyncratic drug reactions. Drug Metab
Rev 24:299-366 (1992).
94. Neumann H-G. Analysis of hemoglobin as a dose monitor
for alkylating and arylating agents. Arch Toxicol 56:1-6 (1984).
95. Neumann H-G. Haemoglobin binding in control of exposure
and risk assessment of aromatic amines. In: IARC Monographs on Methods for
Detecting DNA-damaging Agents in Humans: Application in Cancer Epidemiology
and Prevention, Vol 89 (Bartsch H, Hemminki K, O'Neill ID, eds). Lyon: International
Agency for Research on Cancer, 1989;157-165.
96. Sabbioni G. Hemoglobin binding of monocyclic aromatic
amines: molecular dosimetry and quantitative structure activity relationships
for the N-oxidation. Chem Biol Interact 81:91-117 (1992).
97. Eyer P, Ascherl M. Reactions of para-substituted nitrosobenzenes
with human hemoglobin. Biol Chem Hoppe-Seyler 368:285-294 (1987).
98. Stevens JL, Wallin A. Is the toxicity of cysteine conjugates
formed during mercapturic acid biosynthesis relevant to the toxicity of
covalently bound drug residues? Drug Metab Rev 22:617-635 (1990).
99. Weber N. Overview of bound residue chemistry. Drug
Metab Rev 22:611-615 (1990).
100. Störle C. Aktivierung von N-alkylierten p-Phenylendiaminen
durch Oxyhämoglobin: Bildung und Reaktionen der Wursterschen Radikale.
Thesis. München, Germany:Ludwig-Maximilians-Universität, 1992.
101. Ludwig E, Eckert K-G, Eyer P. Formation of reactive
glutathione S-conjugates during the oxyhemoglobin-mediated sequential oxidation/addition
reactions of 4-dimethylaminophenol. ISSX Proceedings Vol 2:29 (1992).
[
Table
of Contents] [
Citation
in PubMed] [
Related
Articles]
Last Update: October 22, 1998
