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AN884 Tobacco

Extraction of Tobacco-Specific Nitrosamines (TSNAs) from Urine Using ISOLUTE® SLE+ Prior to UPLC/MS/MS Analysis

28 June 2017

This application note from Biotage Labs describes a method for extracting nitrosamines from urine. The ISOLUTE® SLE+ sample preparation method allows for a fast extraction with sufficient sample clean up, resulting in clean extracts.


TSNAs, or tobacco-specific nitrosamines, are carcinogens found in tobacco products, including e-cigarettes and smokeless tobacco. NNK (4 (methylnitrosamino)-1-(3-pyridyl)-1-butanone), NNAL (4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol), and NNN (n-nitrosonornicotine) are the most commonly analyzed TSNAs. NNK has been seen to cause lung cancer in laboratory animals, NNAL is a metabolite of NNK, and NNN has been seen to cause esophageal cancer in laboratory animals. This proof of concept study focusses on NNN (n-nitrosonornicotine).

TSNAs can be difficult to accurately detect, as, like nicotine, there can be false positives for secondhand and third-hand contamination. Secondhand contamination, or secondhand smoke, causes non-smokers to test positive for TSNAs if the detection limits for the method are low. Third-hand contamination can be from test tubes, extraction media, pipette tips, or any number of items used throughout the sample extraction and analysis processes.

Download the Application Note Below »


This proof of concept study focusses on:

  •  NNN (n-nitrosonornicotine)

But the supported liquid extraction methodology described is a also suitable for:

  • NNK (4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone)
  • NNAL (4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol)



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