Long-Term Stability of Volatile Nitrosamines in Human Urine

Volatile nitrosamines (VNAs) are established teratogens and carcinogens in animals and classified as probable (group 2A) and possible (group 2B) carcinogens in humans by the IARC. High levels of VNAs have been detected in tobacco products and in both mainstream and sidestream smoke. VNA exposure may lead to lipid peroxidation and oxidative stress (e.g., inflammation), chronic diseases (e.g., diabetes) and neurodegenerative diseases (e.g., Alzheimer's disease). To conduct epidemiological studies on the effects of VNA exposure, short-term and long-term stabilities of VNAs in the urine matrix are needed. In this report, the stability of six VNAs (N-nitrosodimethylamine, N-nitrosomethylethylamine, N-nitrosodiethylamine, N-nitrosopiperidine, N-nitrosopyrrolidine and N-nitrosomorpholine) in human urine is analyzed for the first time using in vitro blank urine pools fortified with a standard mixture of all six VNAs. Over a 24-day period, analytes were monitored in samples stored at ∼20°C (collection temperature), 4-10°C (transit temperature) and -20 and -70°C (long-term storage temperatures). All six analytes were stable for 24 days at all temperatures (n = 15). The analytes were then analyzed over a longer time period at -70°C; all analytes were stable for up to 1 year (n = 62). A subset of 44 samples was prepared as a single batch and stored at -20°C, the temperature at which prepared samples are stored. These prepared samples were run in duplicate weekly over 10 weeks, and all six analytes were stable over the entire period (n = 22).