Nitration of gamma-tocopherol prevents its oxidative metabolism by HepG2 cells.

Abstract

Gamma-tocopherol (gammaT) is one of the major forms of vitamin E consumed in the diet. Previous reports have suggested increased levels of nitrated gamma-tocopherol (5-NO2-gammaT) in smokers and individuals with conditions associated with elevated nitrative stress. The monitoring of 5-NO2-gammaT and its possible metabolite(s) may be a useful marker of reactive nitrogen species generation in vivo. The major pathway for the metabolism of gammaT is the cytochrome P450 dependent oxidation to its water-soluble metabolite gamma-CEHC, which is excreted in urine. In order to determine if 5-NO2-gammaT could be metabolised via the same route and detected in urine we developed a sensitive gas chromatography-mass spectrometry assay for 5-NO2-gamma-CEHC. 5-NO2-gamma-CEHC was synthesised and its structure confirmed by proton nuclear magnetic resonance and mass spectrometry. While gamma-CEHC was abundant in urine from healthy volunteers, as well as patients with coronary heart disease and type 2 diabetes, 5-NO2-gamma-CEHC was undetectable (limit of detection of 5 nM). To understand this observation we examined the uptake and metabolism of gammaT and 5-NO2-gammaT by HepG2 cells. gammaT was readily incorporated into cells and metabolised to gamma-CEHC over a period of 48 hours. In contrast, 5-NO2-gammaT was poorly incorporated into HepG2 cells and not metabolised to 5-NO2-gamma-CEHC over the same time period. We conclude that nitration of gammaT prevents its incorporation into liver cells and therefore its metabolism to the water-soluble metabolite. Whether 5-NO2-gammaT could be metabolised via other pathways in vivo requires further investigation.

• Authors: Jason H Y Wu, Jonathan M Hodgson, Natalie C Ward, Michael W Clarke, Ian B Puddey, Kevin D Croft
• Journal: Free radical biology & medicine (Free Radic Biol Med) Vol. 39 Issue 4 Pg. 483-94 (Aug 15 2005) ISSN: 0891-5849 [Print] United States
• PMID: 16043020 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't, Validation Study)

Chemical References

• 5-nitro-gamma-tocopherol
• Chromans
• Cytochrome P-450 Enzyme Inhibitors
• Propionates
• Reactive Nitrogen Species
• 2,7,8-trimethyl-2-(beta-carboxyethyl)-6-hydroxychroman
• gamma-Tocopherol
• Cytochrome P-450 Enzyme System

Topics

• Adult
• Carcinoma, Hepatocellular (metabolism)
• Cell Line, Tumor
• Chromans (urine)
• Chromatography, High Pressure Liquid
• Cytochrome P-450 Enzyme Inhibitors
• Cytochrome P-450 Enzyme System (metabolism)
• Gas Chromatography-Mass Spectrometry
• Humans
• Male
• Propionates (urine)
• Reactive Nitrogen Species (metabolism)
• gamma-Tocopherol (analogs & derivatives, metabolism, urine)