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Anal Biochem. 2009 Nov 15;394(2):243-8. doi: 10.1016/j.ab.2009.07.035. Epub 2009 Jul 28.

A gas chromatography-mass spectrometry method for the quantitation of N-nitrosoproline and N-acetyl-S-allylcysteine in human urine: application to a study of the effects of garlic consumption on nitrosation.

Author information

1
Phytonutrients Laboratory, USDA Human Nutrition Research Center, Beltsville, MD 20705, USA.

Abstract

Biomarkers in urine can provide useful information about the bioactivation of chemical carcinogens and can be used to investigate the chemoprotective properties of dietary nutrients. N-Nitrosoproline (NPRO) excretion has been used as an index for endogenous nitrosation. In vitro and animal studies have reported that compounds in garlic may suppress nitrosation and inhibit carcinogenesis. We present a new method for extraction and sensitive detection of both NPRO and N-acetyl-S-allylcysteine from urine. The latter is a metabolite of S-allylcysteine, which is found in garlic. Urine was acidified and the organic acids were extracted by reversed-phase extraction (RP-SPE) and use of a polymeric weak anion exchange (WAX-SPE) resin. NPRO was quantified by isotope dilution gas chromatography-mass spectrometry (GC-MS) using [13C5]NPRO and N-nitrosopipecolinic acid (NPIC) as internal standards. This method was used to analyze urine samples from a study that was designed to test whether garlic supplementation inhibits NPRO synthesis. Using this method, 2.4 to 46.0 ng NPRO/ml urine was detected. The method is straightforward and reliable, and it can be performed with readily available GC-MS instruments. N-Acetyl-S-allylcysteine was quantified in the same fraction and detectable at levels of 4.1 to 176.4 ng/ml urine. The results suggest that 3 to 5 g of garlic supplements inhibited NPRO synthesis to an extent similar to a 0.5-g dose of ascorbic acid or a commercial supplement of aged garlic extract. Urinary NPRO concentration was inversely associated with the N-acetyl-S-allylcysteine concentration. It is possible that allyl sulfur compounds found in garlic may inhibit nitrosation in humans.

PMID:
19643074
PMCID:
PMC2755231
DOI:
10.1016/j.ab.2009.07.035
[Indexed for MEDLINE]
Free PMC Article

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