Limits...
Quantitative analysis of the oxidative DNA lesion, 2,2-diamino-4-(2-deoxy-beta-D-erythro-pentofuranosyl)amino]-5(2H)-oxazolone (oxazolone), in vitro and in vivo by isotope dilution-capillary HPLC-ESI-MS/MS.

Matter B, Malejka-Giganti D, Csallany AS, Tretyakova N - Nucleic Acids Res. (2006)

Bottom Line: While the amounts of oxazolone continued to increase with the duration of irradiation, those of 8-oxo-dG reached a maximum at 20 min, suggesting that 8-oxo-dG is converted to secondary oxidation products.Both lesions were found in rat liver DNA isolated under carefully monitored conditions to minimize artifactual oxidation.The formation of oxazolone lesions in rat liver DNA, their relative stability in the presence of oxidants and their potent mispairing characteristics suggest that oxazolone may play a role in oxidative stress-mediated mutagenesis.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicinal Chemistry, University of Minnesota Cancer Center, University of Minnesota, Minneapolis, MN 55455, USA.

ABSTRACT
A major DNA oxidation product, 2,2-diamino-4-[(2-deoxy-beta-D-erythro-pentofuranosyl)amino]-5(2H)-oxazolone (oxazolone), can be generated either directly by oxidation of dG or as a secondary oxidation product with an intermediate of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxo-dG). Site-specific mutagenesis studies indicate that oxazolone is a strongly mispairing lesion, inducing approximately 10-fold more mutations than 8-oxo-dG. While 8-oxo-dG undergoes facile further oxidation, oxazolone appears to be a stable final product of guanine oxidation, and, if formed in vivo, can potentially serve as a biomarker of DNA damage induced by oxidative stress. In this study, capillary liquid chromatography-electrospray ionization tandem mass spectrometry (HPLC-ESI-MS/MS) methods were developed to enable quantitative analysis of both 8-oxo-dG and oxazolone in DNA from biological sources. Sensitive and specific detection of 8-oxo-dG and oxazolone in enzymatic DNA hydrolysates was achieved by isotope dilution with the corresponding 15N-labeled internal standards. Both nucleobase adducts were formed in a dose-dependent manner in calf thymus DNA subjected to photooxidation in the presence of riboflavin. While the amounts of oxazolone continued to increase with the duration of irradiation, those of 8-oxo-dG reached a maximum at 20 min, suggesting that 8-oxo-dG is converted to secondary oxidation products. Both lesions were found in rat liver DNA isolated under carefully monitored conditions to minimize artifactual oxidation. Liver DNA of diabetic and control rats maintained on a diet high in animal fat contained 2-6 molecules of oxazolone per 10(7) guanines, while 8-oxo-dG amounts in the same samples were between 3 and 8 adducts per 10(6) guanines. The formation of oxazolone lesions in rat liver DNA, their relative stability in the presence of oxidants and their potent mispairing characteristics suggest that oxazolone may play a role in oxidative stress-mediated mutagenesis.

Show MeSH

Related in: MedlinePlus

Chemical structures of the oxidative DNA lesions examined in the present study.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC1636462&req=5

sch1: Chemical structures of the oxidative DNA lesions examined in the present study.

Mentions: Reactive oxygen and nitrogen species, e.g. hydrogen peroxide, hydroxyl radical, superoxide, peroxynitrite and singlet oxygen, are produced in normal tissues as a result of aerobic metabolism, immune response and inflammation (1,2). The oxidative degradation of DNA has been implicated in aging, cancer and in some degenerative diseases (3–6). For example, smokers who exhibit low levels of hOgg1, a repair protein responsible for the removal of oxidative DNA lesions, are at an increased risk of lung cancer (7). Among the four DNA bases, guanine is most susceptible to oxidation because it has the lowest oxidation potential (8,9). Guanine oxidation is further facilitated in the GG and GGG repeats (8,10–12). Guanine oxidation gives rise to a variety of products, including 8-oxo-dG, spiroiminodihydantoin, guanidinohydantoin, 2-amino-5-[2-deoxy-β-d-erythro-pentofuranosyl)amino]-4H-imidazol-4-one (imidazolone) and its hydrolysis product, 2,2-diamino-4-[(2-deoxy-β-d-erythro-pentofuranosyl)amino]-5(2H)-oxazolone (oxazolone) (Scheme 1) (13–19).


Quantitative analysis of the oxidative DNA lesion, 2,2-diamino-4-(2-deoxy-beta-D-erythro-pentofuranosyl)amino]-5(2H)-oxazolone (oxazolone), in vitro and in vivo by isotope dilution-capillary HPLC-ESI-MS/MS.

Matter B, Malejka-Giganti D, Csallany AS, Tretyakova N - Nucleic Acids Res. (2006)

Chemical structures of the oxidative DNA lesions examined in the present study.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC1636462&req=5

sch1: Chemical structures of the oxidative DNA lesions examined in the present study.
Mentions: Reactive oxygen and nitrogen species, e.g. hydrogen peroxide, hydroxyl radical, superoxide, peroxynitrite and singlet oxygen, are produced in normal tissues as a result of aerobic metabolism, immune response and inflammation (1,2). The oxidative degradation of DNA has been implicated in aging, cancer and in some degenerative diseases (3–6). For example, smokers who exhibit low levels of hOgg1, a repair protein responsible for the removal of oxidative DNA lesions, are at an increased risk of lung cancer (7). Among the four DNA bases, guanine is most susceptible to oxidation because it has the lowest oxidation potential (8,9). Guanine oxidation is further facilitated in the GG and GGG repeats (8,10–12). Guanine oxidation gives rise to a variety of products, including 8-oxo-dG, spiroiminodihydantoin, guanidinohydantoin, 2-amino-5-[2-deoxy-β-d-erythro-pentofuranosyl)amino]-4H-imidazol-4-one (imidazolone) and its hydrolysis product, 2,2-diamino-4-[(2-deoxy-β-d-erythro-pentofuranosyl)amino]-5(2H)-oxazolone (oxazolone) (Scheme 1) (13–19).

Bottom Line: While the amounts of oxazolone continued to increase with the duration of irradiation, those of 8-oxo-dG reached a maximum at 20 min, suggesting that 8-oxo-dG is converted to secondary oxidation products.Both lesions were found in rat liver DNA isolated under carefully monitored conditions to minimize artifactual oxidation.The formation of oxazolone lesions in rat liver DNA, their relative stability in the presence of oxidants and their potent mispairing characteristics suggest that oxazolone may play a role in oxidative stress-mediated mutagenesis.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicinal Chemistry, University of Minnesota Cancer Center, University of Minnesota, Minneapolis, MN 55455, USA.

ABSTRACT
A major DNA oxidation product, 2,2-diamino-4-[(2-deoxy-beta-D-erythro-pentofuranosyl)amino]-5(2H)-oxazolone (oxazolone), can be generated either directly by oxidation of dG or as a secondary oxidation product with an intermediate of 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxo-dG). Site-specific mutagenesis studies indicate that oxazolone is a strongly mispairing lesion, inducing approximately 10-fold more mutations than 8-oxo-dG. While 8-oxo-dG undergoes facile further oxidation, oxazolone appears to be a stable final product of guanine oxidation, and, if formed in vivo, can potentially serve as a biomarker of DNA damage induced by oxidative stress. In this study, capillary liquid chromatography-electrospray ionization tandem mass spectrometry (HPLC-ESI-MS/MS) methods were developed to enable quantitative analysis of both 8-oxo-dG and oxazolone in DNA from biological sources. Sensitive and specific detection of 8-oxo-dG and oxazolone in enzymatic DNA hydrolysates was achieved by isotope dilution with the corresponding 15N-labeled internal standards. Both nucleobase adducts were formed in a dose-dependent manner in calf thymus DNA subjected to photooxidation in the presence of riboflavin. While the amounts of oxazolone continued to increase with the duration of irradiation, those of 8-oxo-dG reached a maximum at 20 min, suggesting that 8-oxo-dG is converted to secondary oxidation products. Both lesions were found in rat liver DNA isolated under carefully monitored conditions to minimize artifactual oxidation. Liver DNA of diabetic and control rats maintained on a diet high in animal fat contained 2-6 molecules of oxazolone per 10(7) guanines, while 8-oxo-dG amounts in the same samples were between 3 and 8 adducts per 10(6) guanines. The formation of oxazolone lesions in rat liver DNA, their relative stability in the presence of oxidants and their potent mispairing characteristics suggest that oxazolone may play a role in oxidative stress-mediated mutagenesis.

Show MeSH
Related in: MedlinePlus