Acrolein Mercapturates: Synthesis, Characterization, and Assessment of Their Role in the Bladder Toxicity of Cyclophosphamide

Ramu, Kumar; Fraiser, Lucy; Mamiya, Blain M.; Ahmed, Tamer; Kehrer, James P.

doi:10.1021/tx00046a005

Cited by 64 publications

(54 citation statements)

References 31 publications

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“…20 ILL of acrolein (0.30 mmo1) was added by micropipette. 1 H NMR results are in agreement with spectra published earlier [37,40] …”

supporting

confidence: 91%

“…Acrolein thioether conjugates were chosen as model compounds for Michael-thioether conjugates. Acrolein is an important industrial chemical [1,361, a metabolite of the cancer chemotherapeutic agent cyclophosphamide, [36][37][38] and an unwanted side-product of combustion and cigarette smoke [38,39]. Compounds for this study are acrolein conjugates of N-acetyl-L-cysteine, glutathione, and N-acetylglutathione.…”

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confidence: 99%

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Fragmentation of protonated thioether conjugates of acrolein using low collision energies

Oberth

Jones

1997

J. Am. Soc. Mass Spectrom.

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The protonated mercapturic acid conjugate of acrolein, S-(3-oxopropyI)-N-acetyl-L-cysteine (I), undergoes facile retro-Michael loss of acrol ein in the gas phase. To determine whether extensive loss of acrolein would impede structural characterization of acrolein-peptide ad ducts, fragmentation reactions of a series of conjugates, formed by l,4-Michael addition of acrolein to peptides and cysteine derivatives, w ere investigated at collision cell potentials up to -50 V using a triple quadrupole mass spectrometer. Differences in fragmentation dynamics suggest protonation at the sulfur of the N-ac etylcysteine conjugate 1 facilitates retro-Michael elimination of acrolein with a low activation energy relative to other fragmentations. Analogous fragmentation was eliminated after borohydride reduction of the aldehyde to an alcohol. Retro-Michael fragmentation was not significant for acrolein conjugates of glutathione derivatives, suggesting that proton sequestration occurs in peptides with multiple amide linkages even when the peptide does not contain a basic amino group. An unexpected outcome of these experiments was the observation of a facile gas-phase cleavage of peptides on the N-terminal side of S-(3-oxopropylkysteine residues. Such fragmentation behavior may prove useful for locating cysteine residues in peptides. . Protein sulfhydryl groups are especially vulnerable to electrophiIic attack and form thioether conjugates whose presence indicates mechanisms of metabolic activation or detoxification. Knowledge of protein targets and sites of covalent attachment can be used to identify reactive metabolites, and these modified proteins can serve as biomarkers of exposure to xenobiotics in clinical and epidemiological studies [1][2][3][4][5][6][7][8]. One important class of xenobiotic-protein adducts forms via lA-Michael addition of a ,,B-unsaturated carbonyls to nucleophilic amino acid residues. Many a,,B-unsaturated carbonyls including acrylic monomers, combustion by-products or metabolites such as acrolein [1], and endogenous compounds such as 4-hydroxynonenal react with protein nucleophilic side chains via Michael addition [9][10][11].Mass spectrometry has emerged as a powerful analytical tool for the characterization of post-translationally modified peptides and proteins because this approach can identify the modifying group and sites of modification [12]. Earlier studies demonstrated the utility of fast atom bombardment mass spectrometry (FAB/MS) for identification of specific thioether conjuAddress reprint requests to Dr. A. Daniel Jones, Facility for Adv anced Instrumen tation, University of California, Dav is, CA 95616. E-mail: adjones@ucdavis.edu gates from biological samples [3,4,[13][14][15][16][17] . Using FAB or newer ionization techniques coupled with tandem mass spectrometry (MS/MS), researchers determined structures of thioether conjugates extracted from biological matrices [3][4][5][6][7][8][13][14][15][16][17][18][19][20][21][22]. In these MS/MS experiments, structural information was obtained fro...

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“…20 ILL of acrolein (0.30 mmo1) was added by micropipette. 1 H NMR results are in agreement with spectra published earlier [37,40] …”

supporting

confidence: 91%

mentioning

confidence: 99%

Fragmentation of protonated thioether conjugates of acrolein using low collision energies

Oberth

Jones

1997

J. Am. Soc. Mass Spectrom.

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“…This adduct was synthesized according to the N-acetyl-3-oxopropyl-L-cysteine synthesis of Ramu et al [23] using acrylamide in place of acrolein and characterized by comparison of recently published mass spectral data [24].…”

Section: Synthesis Ofmentioning

confidence: 99%

Rapid detection and identification ofN-acetyl-L-cysteine thioethers using constant neutral loss and theoretical multiple reaction monitoring combined with enhanced product-ion scans on a linear ion trap mass spectrometer

Scholz

Dekant

Völkel

et al. 2005

J. Am. Soc. Mass Spectrom.

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A sensitive and specific liquid chromatography-mass spectrometry (LC-MS) method based on the combination of constant neutral loss scans (CNL) with product ion scans was developed on a linear ion trap. The method is applicable for the detection and identification of analytes with identical chemical substructures (such as conjugates of xenobiotics formed in biological systems) which give common CNLs. A specific CNL was observed for thioethers of N-acetyl-L-cysteine (mercapturic acids, MA) by LC-MS/MS. MS and HPLC parameters were optimized with 16 MAs available as reference compounds. All of these provided a CNL of 129 Da in the negative-ion mode. To assess sensitivity, a multiple reaction monitoring (MRM) mode with 251 theoretical transitions using the CNL of 129 Da combined with a product ion scan (IDA thMRM) was compared with CNL combined with a product ion scan (IDA CNL). An information-dependent acquisition (IDA) uses a survey scan such as MRM (multiple reaction monitoring) to generate "informations" and starting a second acquisition experiment such as a product ion scan using these "informations." Th-MRM means calculated transitions and not transitions generated from an available standard in the tuning mode. The product ion spectra provide additional information on the chemical structure of the unknown analytes. All MA standards were spiked in low concentrations to rat urines and were detected with both methods with LODs ranging from 60 pmol/mL to 1.63 nmol/mL with IDA thMRM. The expected product ion spectra were observed in urine. Application of this screening method to biological samples indicated the presence of a number of MAs in urine of unexposed rats, and resulted in the identification of 1,4-dihydroxynonene mercapturic acid as one of these MAs by negative and positive product ion spectra. These results show that the developed methods have a high potential to serve as both a prescreen to detect unknown MAs and to identify these analytes in complex matrix. (J Am Soc Mass Spectrom 2005, 16, 1976 -1984

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“…In previous studies, both oxidized and reduced forms of mercapturic acid derivatives of HNE and acrolein have been shown (42,43). However, it is not clear whether these redox transformations occur with the GS-X, X-Cys-Gly, Cys-X, or N-acetyl-Cys-X.…”

mentioning

confidence: 95%

“…Studies with mercapturic acids of acrolein show that in contrast to the oxidized and untransformed mercapturates, the reduced mercapturic acid derivative is not toxic (43), suggesting that reduction may represent a metabolic transformation to abrogate the toxicity of GSH conjugates. While the GSH conjugates are covalent adducts, several of these can spontaneously dissociate.…”

mentioning

confidence: 99%

Metabolism of the Lipid Peroxidation Product, 4-Hydroxy-trans-2-nonenal, in Isolated Perfused Rat Heart

Srivastava¹,

Chandra²,

Wang³

et al. 1998

Journal of Biological Chemistry

221

176

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The metabolism of 4-hydroxy-trans-2-nonenal (HNE), an ␣,␤-unsaturated aldehyde generated during lipid peroxidation, was studied in isolated perfused rat hearts. High performance liquid chromatography separation of radioactive metabolites recovered from [ 3 H]HNE-treated hearts revealed four major peaks. Based on the retention times of synthesized standards, peak I, which accounted for 20% radioactivity administered to the heart, was identified to be due to glutathione conjugates of HNE. Peaks II and III, containing 2 and 37% radioactivity, were assigned to 1,4-dihydroxy-2-nonene (DHN) and 4-hydroxy-2-nonenoic acid, respectively. Peak IV was due to unmetabolized HNE. The electrospray ionization mass spectrum of peak I revealed two prominent metabolites with m/z values corresponding to [M ؉ H] ؉ of HNE and DHN conjugates with glutathione. The presence of 4-hydroxy-2-nonenoic acid in peak III was substantiated using gas chromatography-chemical ionization mass spectroscopy. When exposed to sorbinil, an inhibitor of aldose reductase, no GS-DHN was recovered in the coronary effluent, and treatment with cyanamide, an inhibitor of aldehyde dehydrogenase, attenuated 4-hydroxy-2-nonenoic acid formation. These results show that the major metabolic transformations of HNE in rat heart involve conjugation with glutathione and oxidation to 4-hydroxy-2-nonenoic acid. Further metabolism of the GS-HNE conjugate involves aldose reductasemediated reduction, a reaction catalyzed in vitro by homogenous cardiac aldose reductase.

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Acrolein Mercapturates: Synthesis, Characterization, and Assessment of Their Role in the Bladder Toxicity of Cyclophosphamide

Cited by 64 publications

References 31 publications

Fragmentation of protonated thioether conjugates of acrolein using low collision energies

Fragmentation of protonated thioether conjugates of acrolein using low collision energies

Rapid detection and identification ofN-acetyl-L-cysteine thioethers using constant neutral loss and theoretical multiple reaction monitoring combined with enhanced product-ion scans on a linear ion trap mass spectrometer

Metabolism of the Lipid Peroxidation Product, 4-Hydroxy-trans-2-nonenal, in Isolated Perfused Rat Heart

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