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R N Loeppky

Publications and source records attributed to R N Loeppky.

At least 19 recordsLinked to original sources
PubMed2000

N-Alkyl-N-cyclopropylanilines as mechanistic probes in the nitrosation of N,N-dialkyl aromatic amines.

A group of N-cyclopropyl-N-alkylanilines has been synthesized, and their reaction with nitrous acid in aqueous acetic acid at 0 degrees C was examined. All compounds reacted rapidly to produce the corresponding N-alkyl-N-nitrosoaniline by specific cleavage of the cyclopropyl group from the nitrogen. The transformations were unaffected by the nature of the alkyl substituent (Me, Et, (i)()Pr, Bn). The reaction of 4-chloro-N-2-phenylcyclopropyl-N-methylaniline with nitrous acid gave 4-chloro-N-methyl-N-nitrosoaniline (76%), cinnamaldehyde (55%), 3-phenyl-5-hydroxyisoxazoline (26%), and 5-(N-4-chlorophenylmethylamino)-3-phenylisoxazoline (8%). Both the selective cleavage of the cyclopropyl group from the aromatic amine nitrogen and nature of the products derived from the cyclopropane ring support a mechanism involving the formation of an amine radical cation. This step is followed by rapid cyclopropyl ring opening to produce an iminium ion with a C-centered radical which either combines with NO or is oxidized.

Amines↗
PubMed2000

In vitro DNA deamination by alpha-nitrosaminoaldehydes determined by GC/MS-SIM quantitation.

The deamination of DNA bases by three alpha-nitrosaminoaldehydes, butylethanalnitrosamine, methylethanalnitrosamine, and N-nitroso-2-hydroxymorpholine (NHMOR), the direct metabolite of potent animal carcinogen N-nitrosodiethanolamine, was demonstrated by a set of in vitro experiments. The deamination of guanine, adenine, and cytosine bases in nucleotides, oligonucleotides, and calf thymus DNA gave xanthine, hypoxanthine, and uracil, respectively. The order of relative reactivities of the bases was as listed above. Deamination of cytosine to uracil was detected by the reaction of (32)P-labeled oligonucleotide ([5'-(32)P]CGAT) followed by enzymatic hydrolysis. Quantitative analysis of deamination of guanine and adenine in calf thymus DNA was performed by a gas chromatography/mass spectrometry-selected ion monitoring method. Both the extent and the rate of the deamination reactions which occur by transnitrosation from the alpha-nitrosaminoaldehyde to the base were determined for formation of xanthine and hypoxanthine. The deamination of guanine by NHMOR remained significant at low substrate levels.

Adenine↗
PubMed1999

Glyoxal-guanine DNA adducts: detection, stability and formation in vivo from nitrosamines.

The glyoxal-deoxyguanosine adduct (gG) is formed from alpha-nitrosamino aldehydes and dG in vitro and in vivo from nitrosamines carrying the 2-hydroxyethyl side-chain as well as from N-nitrosomorpholine. The structures of all of the diastereomeric forms of both the cis and trans isomers of the adduct have been investigated by ab initio calculations and with nuclear magnetic resonance spectroscopy at 500 MHz. The preferred orientation of the OH groups is trans, but at equilibrium a small amount of the cis isomer was observed. The pH-independent equilibrium constant for the hydrolysis of the gG adduct is K = 1.36 x 10(-4) mol/L, and its rate of formation at pH 7.3 is k = 5.3 min-1 mol-1. In acid (pH 2), the hydrolysis of the nucleosidic linkage is nearly twice as rapid as the hydrolysis of gG to glyoxal and dG. We used a gG analogue to explore a number of reductive methods for derivatization of the adduct, but all of the processes either gave low yields or product mixtures which rendered them impractical for derivatizing the adduct in DNA. A 32P-postlabelling method for detection of the pH-sensitive gG adduct has been developed, which permitted detection of the adduct in the liver DNA of male Wistar rats after administration of selected nitrosamines. The levels of adducts found were: N-nitrosodiethanolamine > 2-hydroxyethylmethynitrosamine > N-nitrosomorpholine > 2-hydroxyethyethylnitrosamine. In separate experiments, N-nitrosodiethanolamine gave greater adduct levels than its metabolite 2-hydroxy-N-nitrosomorpholine. Mechanistic pathways for the generation of gG adducts in vitro and in vivo are discussed.

Animals↗
PubMed1998

Probing the mechanism of the carcinogenic activation of N-nitrosodiethanolamine with deuterium isotope effects: in vivo induction of DNA single-strand breaks and related in vitro assays.

A series of bioassays, including in vivo induction of DNA single-strand breaks (SSB) and cytotoxicity in cytochrome P450 2E1-transfected cells, were utilized with N-nitrosodiethanolamine (NDELA), its deuterated isotopomers (alpha-D4NDELA and beta-D4NDELA), N-nitroso-2-hydroxymorpholine (NHMOR), and two of its deuterated isotopomers (2-D-NHMOR and 5,5-D2-NHMOR) to probe the mechanism of carcinogenic activation of NDELA and the role of its metabolite NHMOR. DNA samples, taken from the livers of male Wistar rats 4 h after the administration of NDELA, exhibited dose-dependent DNA SSB levels over the range of 0.08-0.75 mmol/kg (body weight), with the greatest SSB level at the highest dose. Deuterium isotope effects on DNA SSB levels were inversely dependent on dose: alpha-D4NDELA, 3. 22-1.37; and beta-D4NDELA, 1.38-0.79. At the lowest dose of 0.15 mmol/kg (body weight), 5,5-D2-NHMOR gave an isotope effect for DNA SSB of 2.8 while that for 2-D-NHMOR was 0.7. NDELA and beta-D4NDELA were equally cytotoxic to human P450 2E1-transfected V79 Chinese hamster cells, while alpha-D4NDELA was not. Significant DNA SSB levels were observed in these cells for NDELA and beta-D4NDELA but not for alpha-D4NDELA. A kinetic deuterium isotope effect of 2.6 for Vmax/Km was observed for the horse liver alcohol dehydrogenase-mediated oxidation of beta-D4NDELA to NHMOR, while kH/kD for alpha-D4NDELA was 1.05. These data provide the first definitive evidence for the activation of NDELA by a pathway involving the scission of the alpha-CH bond and are consistent with P450 2E1-mediated alpha-hydroxylation of NDELA producing the corresponding reactive alpha-hydroxynitrosamine.

Animals↗
PubMed1995

Thiol oxidation by 1,2,3-oxadiazolinium ions, presumed carcinogens.

3-Alkyl-1,2,3-oxadiazolinium ions 1 have been proposed as reactive intermediates in the activation of (2-hydroxyethyl)nitrosamines. The reaction of 3-methyl-1,2,3-oxadiazolinium tosylate (1a), 2-ethyl-1-methoxy-2-phenyldiazenium tetrafluoroborate (3), and 3-phenyl-1,2,3-oxadiazolinium triflate (1b) with thiols was investigated to determine the behavior of these compounds toward typical "cellular nucleophiles". Each of these substances oxidizes benzenethiol to diphenyl disulfide. The reaction in aqueous buffer at pH 7.4 is rapid. Reaction of 1b with benzenethiol gives, in addition to the disulfide, benzene, biphenyl, azobenzene, diphenyl sulfide, aniline, and glycolaldehyde. Similar products are obtained from 3. Phenyldiazene is postulated as an intermediate in this process, and its generation from phenyldiazoformate in the presence of benzenethiol gives similar products. Diazenes are presumed to arise by proton abstraction from the CH adjacent to N. The kinetics of reaction of 1a with N-acetylcysteine to give the corresponding disulfide show first order dependence on each reactant and base catalysis. The data from these model chemical experiments suggest that 1,2,3-oxadiazolinium ions could react with abundant thiols in cells to lead to either their detoxification or radical processes emanating from diazenes. The occurrence of thiol-oxadiazolinium ion redox transformations could modulate the alkylation chemistry of these substances as well.

Carcinogens↗
PubMed1991

Nitrosation of tertiary aromatic amines related to sunscreen ingredients.

Possible routes to the formation of the sunscreen contaminant, 2-ethylhexyl 4-N-methyl-N-nitrosoaminobenzoate, have been investigated in a study of the nitrosation chemistry of 2-ethylhexyl 4-N,N-dimethylaminobenzoate (Padimate-O) and related tertiary and secondary amines. Padimate-O and the corresponding ethyl ester nitrosate rapidly at 25 degrees C in either N2O3:ether or HNO2:HOAc to produce a mixture of alkyl 4-N-methyl-N-nitrosoaminobenzoate and alkyl 4-N,N-dimethylamino-3-nitrobenzoate, the former of which is the major product. The nitrosative dealkylation of these amines at this low temperature is unusual. Asymmetrical amines exhibit a preference for nitrosative demethylation (methyl versus ethyl or benzyl), but the cleavage ratios in N2O3:ether are time-dependent, suggesting competing mechanisms with different reactant kinetic orders. A radical cation route would explain the unusual reactivity, which may compete with the established nitrosative dealkylation mechanism. 2-Ethylhexyl 4-N-methyl-N-nitrosoaminobenzoate was mutagenic in two strains of Salmonella typhimurium in the Ames assay.

4-Aminobenzoic Acid↗
PubMed1991

Mutagenicity, DNA damage and DNA adduct formation by N-nitroso-2-hydroxyalkylamine and corresponding aldehydes.

The potent carcinogen N-nitrosodiethanolamine (NDELA) becomes mutagenic to Salmonella typhimurium TA98 and TA100 when activated by alcohol dehydrogenase from yeast or horse liver. Metabolic pathways different from alpha-oxidation might therefore be important for the activation of N-nitroso-2-hydroxyalkylamines such as NDELA. In an in-vitro test system (Namalva cells), neither NDELA nor N-nitrosoethyl-2-hydroxyethylamine was genotoxic, whereas the corresponding metabolites from alcohol dehydrogenase-mediated oxidation, N-nitroso-2-hydroxymorpholine and N-nitrosoethylethanalamine, induced single-strand breaks even at low doses. An immuno-slot-blot assay was used to study the formation of O6-2-hydroxyethyldeoxyguanosine in rat liver after oral administration of different N-nitroso-2-hydroxyalkylamines. When given at equimolar doses (0.375 mmol/kg), DNA hydroxyethylation was considerably lower (6.7 mumol/mol deoxyguanosine) with NDELA than with N-nitrosoethyl-2-hydroxyethylamine (48.7 mumol/mol deoxyguanosine) or N-nitrosomethyl-2-hydroxyethylamine (72.1 mumol/mol deoxyguanosine). N-Nitroso-2-hydroxymorpholine did not form detectable levels of O6-2-hydroxyethyldeoxyguanosine.

Aldehydes↗
PubMed1991

Nitrosamine activation and detoxication through free radicals and their derived cations.

The enzymatic activation of simple dialkylnitrosamines is widely perceived to involve cytochrome P450-mediated alpha-hydroxylation to generate an unstable alpha-hydroxynitrosamine. This process can also result in the denitrosation of the nitrosamine, presumably through a common intermediate. We present evidence that the critical intermediate is the alkynitrosaminomethyl free radical, which we generated by thermal decomposition of a nitrosamino acid perester. The radical rapidly loses NO to generate an N-alkylmethyleneimine. The radical is also produced during the Ce(IV) oxidation of beta-hydroxynitrosamines after fragmentation, where it not only loses NO but is oxidized further to a cation which reacts with water to form an alpha-hydroxynitrosamine. These results provide models of the activation and detoxication pathways for beta-oxidized nitrosamines.

Biotransformation↗
PubMed1987

Nitroso transfer from alpha-nitrosamino aldehydes: implications for carcinogenesis.

Six alkylnitrosamino ethanols (R-N(NO)-CH2CH2OH; R = Me, nBu, sBu, iBu, tBu, HOCH2CH2-), including the potent carcinogen N-nitrosodiethanolamine, have been shown to undergo efficient liver alcohol dehydrogenase catalyzed oxidation to their corresponding alpha-nitrosamino aldehydes. Five structurally representative nitrosamino-ethanals (R-N(NO)CH2CHO, R = 4-ClC6H4-, CH3-, nBu-, tBu-, HOCH2CH2-) have been synthesized. Each of these compounds demonstrates the unusual property of facile transnitrosation to a secondary amine. Transnitrosation to dimethylamine, pyrrolidine, morpholine and N-methylaniline has been shown. This reaction occurs rapidly at room temperature in organic solvents but is somewhat slower in aqueous buffer due to extensive equilibrium formation of gem diols by hydration of the aldehyde group. In aqueous media the transnitrosation rate increases with increasing pH from 7 to 9 and does not occur at pH 4. Transnitrosation to primary amines results in deamination (benzylamine----benzyl alcohol). The transnitrosation reaction is accompanied by the formation of imines of glyoxal (R - N = CH - CH = N - R) which appear as primary amines and glyoxal in aqueous solution. Other products have also been characterized as well. These chemical and biochemical data, taken together with results in other laboratories, provide strong support for our hypothesis that certain beta-oxidized nitrosamines can be activated to proximate or ultimate carcinogens by biochemical oxidation to produce highly reactive nitrosamines.

Aldehydes↗
PubMed1987

Alpha-nitrosaminoaldehydes: highly reactive metabolites.

alpha-Nitrosamino aldehydes are highly reactive compounds which are directly-acting mutagens and are capable of facile transnitrosation to secondary and primary amines. The latter lead reactions to deamination. N-Nitrosobutyl(2-oxoethyl)amine (NBOEA) undergoes spontaneous decomposition in buffer at pH greater than 7 (25 degrees C) to give glyoxal and products implicating the formation of the butyl diazonium ion. NBOEA reacts with guanosine to produce xanthosine (by deamination), 7-butylguanosine and the 1,N2 glyoxal adduct, among other products. All beta-nitrosaminoethanols investigated undergo liver alcohol dehydrogenase-catalysed oxidation to their corresponding aldehydes. Several of these aldehydes have been shown to be directly-acting mutagens. These data provide strong evidence for an alternative carcinogenic bioactivation route for nitrosamines which does not involve alpha-oxidation.

Aldehydes↗
PubMed1984

Ester-mediated nitrosamine formation from nitrite and secondary or tertiary amines.

N-Nitrosamines are formed from the heating of either a secondary or a tertiary amine with sodium nitrite in the presence of a high-boiling ester such as 2-acetoxyethanol in ethylene glycol. The four secondary and six tertiary amines examined were found to produce N-nitrosamines in yields ranging from 4% to 80% when equimolar amounts of amine and ester were heated at 120 degrees C with one- to ten-fold equivalents of sodium nitrite in ethylene glycol. Secondary amines competitively produced acetamides at a rate slightly greater than N-nitrosamine formation. Preincubation of a large excess of sodium nitrite and ester led to the rapid formation of N-nitrosamines in high yield. The reaction of tribenzylamine resulted in the formation of both benzaldehyde and dibenzylnitrosamine. N,N-Dimethylbenzylamine reacted to give nearly equimolar amounts of N-nitrosodimethylamine and N-nitroso-N-methylbenzylamine. It is proposed that the nitrosating agent is a nitrous ester, and it is shown that 2-benzoxyethyl nitrite rapidly nitrosates secondary and tertiary amines under these reaction conditions. It is also proposed that these transformations are good models for the environmental formation of N-nitrosamines in foods and commercial products.

Amines↗
PubMed1984

Alternative bioactivation routes for beta-hydroxynitrosamines. Biochemical and chemical model studies.

The biochemical retroaldol-like fragmentation of beta-hydroxynitrosamines has been investigated further. The extent of fragmentation of 2-hydroxy-2-methylpropyl-methylnitrosamine (HMPMN) to N-nitrosodimethylamine and acetone induced by metabolic activation increases as the NADPH level is decreased. 2-Hydroxy-2-phenylethyl-methylnitrosamine (HPhEMN) undergoes competitive oxidation to 2-oxy-2-phenylethyl-methylnitrosamine (OPhEMN) and fragmentation to benzaldehyde and N-nitroso-dimethylamine in the presence of a metabolic activation system from rat liver. The extent of the oxidation was increased by preinduction of the rats with phenobarbital, or separate addition of NADPH and NAD, but was decreased by addition of dimethyl sulfoxide. The fragmentation was observed most readily when oxidation was inhibited or was not induced by cofactors. When HPhEMN was administered to a rat intraperitoneally, benzaldehyde (fragmentation) was found in the urine with OPhEMN and the substrate, but only the last two substances were found in liver and blood. These experiments provide evidence for retroaldol-like fragmentation of beta-hydroxynitrosamines both in vitro and in vivo. In a related investigation, it was found that N-nitroso-N-4-chlorophenyl-2-aminoethanal (NCAE) is extremely reactive and induces spontaneous generation of 4-chlorobenzenediazonium ion in chloroform, as trapped by 2-naphthol. NCAE reacts with dimethylamine in chloroform, benzene or methanol to give N-nitrosodimethylamine and 4-chloroaniline, among other products. This suggests that beta-nitrosaminoaldehydes produced by the biooxidation of their corresponding alcohols could produce cell alteration through alkylation, deamination or transnitrosation.

Animals↗
PubMed1983

Reducing nitrosamine contamination in cutting fluids.

In simulated metalworking coolants that contained both nitrite and di- or triethanolamine at pH 9, N-nitrosodiethanolamine formed at an initial rate of 11 or 6 ppm/wk, respectively. This rate was increased on heating the fluids, on acidification or by the addition of paraformaldehyde, 1,3,5-trimethylhexahydro-s-triazine, ferricyanide or ferric ethylenediaminetetraacetate. N-Nitrosodiethanolamine also formed when nitrite-free coolants containing either of the two amines above were exposed to nitric oxide in air. No nitrosamines were detected in fluids containing primary amines in place of the secondary and tertiary amines, except that N-nitrosooxazolidine was formed in the fluid containing monoethanolamine after addition of formaldehyde-releasing agents, and N-nitrosodiethanolamine and N-nitrosomorpholine were found in fluid containing diglycolamine (HOCH2CH2OCH2CH2NH2) after the fluid was heated at 100 degrees C for 48 hr. These data suggest several steps by which nitrosamine formation in commercial cutting fluids might be substantially reduced: avoiding acid-splitting as a disposal procedure; removing nitrite from the fluid and/or scavenging adventitious nitrosating agents; avoiding unnecessary heating; adding preservatives to the diluted fluid rather than to the commercial concentrate; replacing inherently nitrosatable amine additives by substitutes which are resistant to nitrosamine formation; minimizing concentrations of catalytically active metal complexes.

Catalysis↗
PubMed1982

Nitrosamine formation from ternary nitrogen compounds.

When amides of pyrrolidine are heated with sodium nitrite, small amounts of NPYR are formed. Both the rate of reaction and the nitrosamine yield are increased when ethylene glycol and glycerine are used as solvents. Kinetic evidence suggests that these polyols react with the amide to form esters and the corresponding amine. The esters then react with sodium nitrite to give nitrite esters which rapidly nitrosate the amine. The heating of LDEA with sodium nitrite gives relatively high yields of NDELA in relatively short times and this nitrosation can be explained by this mechanistic hypothesis. 2-N,N-dimethylaminomethylpyrrole reacts very rapidly with nitrous acid at 25 degrees to give NDMA as the sole nitrosamine. This suggests a new mechanism of tertiary amine nitrosation.

Amides↗