Protocol - NNAL in Urine
Description
NNAL can be measured in urine of people using tobacco products and in urine of non-smokers exposed to secondhand smoke using liquid chromatography - tandem mass spectrometry (LC-MS/MS).
Specific Instructions
Because of the sensitive nature of this assay, analysts cannot be actively using tobacco products, and measurements must be performed in a smoke-free environment.
Safety Precautions: Personal protective equipment (PPE) including eye protection, gloves and suitable protective clothing when used to extract and process samples.
Availability
Protocol
Collect urine sample in a sterile container. Aliquot 10 mL urine sample to be frozen and stored immediately at -20° C.
Liquid chromatography – tandem mass spectrometry (LC-MS/MS) is the preferred method to accurately measure NNAL in urine. (See source references for details.) The limit of quantitation is 0.25 pg/mL.
Personnel and Training Required
Laboratory training in the use of liquid chromatography and tandem mass spectrometry is required, as well as general laboratory training on the safe use of chemicals and solvents.
Equipment Needs
This method requires high-performance liquid chromatography and tandem mass spectrometry on a triple quadrupole mass spectrometer.
Requirements
Requirement Category | Required |
---|---|
Major equipment | Yes |
Specialized training | Yes |
Specialized requirements for biospecimen collection | Yes |
Average time of greater than 15 minutes in an unaffected individual | No |
Mode of Administration
Bioassay
Lifestage
Infant, Toddler, Child, Adolescent, Adult, Senior, Pregnancy
Participants
All participants who can provide urine sample.
Selection Rationale
4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) is a major metabolite of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), the most potent pulmonary carcinogen of all tobacco-specific n-nitrosamines (TSNA). TSNAs are known to be carcinogenic to many animal species and are believed to be carcinogenic to humans as well. These carcinogens are of special significance because they combine an inherent potent pulmonary carcinogenic potential with a high degree of tobacco-exposure specificity because they can be found only in tobacco and tobacco smoke. Measuring NNAL in urine is a reliable way to determine exposure to NNK for smokers, for nonsmokers exposed to environmental tobacco smoke (ETS), and for people who use smokeless tobacco products (e.g., chewing tobacco, snuff).
Language
English
Standards
Standard | Name | ID | Source |
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Derived Variables
None
Process and Review
The Tobacco Regulatory Research (TRR) Content Expert Panel (CEP) reviewed the measures in the Tobacco Regulatory Research collection in February 2024.
Guidance from the TRR CEP includes:
- Replaced protocol
- New Data Dictionary
Previous version in Toolkit archive (link)
Protocol Name from Source
Determination of NNAL in urine using liquid chromatography - tandem mass spectrometry (LC-MS/MS)
Source
Xia Y, McGuffey JE, Bhattacharyya S, Sellergren B, Yilmaz E, Wang L, Bernert JT. Analysis of the tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol in urine by extraction on a molecularly imprinted polymer column and liquid chromatography/atmospheric pressure ionization tandem mass spectrometry. Anal Chem. 2005 Dec 1;77(23):7639-45. doi: 10.1021/ac058027u.PMID: 16316171
Jacob P 3rd, Havel C, Lee DH, Yu L, Eisner MD, Benowitz NL. Subpicogram per milliliter determination of the tobacco-specific carcinogen metabolite 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol in human urine using liquid chromatography-tandem mass spectrometry. Anal Chem. 2008 Nov 1;80(21):8115-21. doi: 10.1021/ac8009005. Epub 2008 Oct 8. PMID: 18841944, PMC3167662
General References
Hecht, S. S. (2002). Human urinary carcinogen metabolites: Biomarkers for investigating tobacco and cancer. Carcinogenesis, 23, 907-922.
Hecht, S. S. (1999). Tobacco smoke carcinogens and lung cancer. Journal of the National Cancer Institute, 91, 1194-1210.
Hecht, S. S., & Hoffmann, D. (1988). Tobacco-specific nitrosamines, an important group of carcinogens in tobacco and tobacco smoke. Carcinogenesis, 9, 875-884.
Preston-Martin, S. (1987). N-nitroso compounds as a cause of human cancer. IARC Scientific Publications, 84, 477-484.
Carmella, S. G., Akerkar, S., & Hecht, S. S. (1993). Metabolites of the tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone in smokers’ urine. Cancer Research, 53, 721-724.
Carmella, S. G., Akerkar, S. A., Richie, J. P., Jr., & Hecht, S. S. (1995). Intraindividual and interindividual differences in metabolites of the tobacco-specific lung carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) in smokers’ urine. Cancer Epidemiology, Biomarkers & Prevention, 4, 635-642.
Carmella, S. G., Yoder, A., & Hecht, S. S. (2006). Combined analysis of r-1,t-2,3,c-4-tetrahydroxy-1,2,3,4-tetrahydrophenanthrene and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol in smokers’ plasma. Cancer Epidemiology, Biomarkers & Prevention, 15, 1490-1494.
Xia, Y., McGuffey, J. E., Bhattacharyya, S., Sellergren, B., Yilmaz, E., Wang, L., & Bernert, J. T. (2005). Analysis of the tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol in urine by extraction on a molecularly imprinted polymer column and liquid chromatography/atmospheric pressure ionization tandem mass spectrometry. Analytical Chemistry, 77, 7639-7645.
Xia, Y., & Bernert, J. T. (in press). Stability of the tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) in urine samples stored at various temperatures. Journal of Analytical Toxicology.
Ashley, D. L., O’Connor, R. J., Bernert, J. T., Watson, C. H., Polzin, G. M., Jain, R. B., et al. (2010). Effect of differing levels of tobacco-specific nitrosamines in cigarette smoke on the levels of biomarkers in smokers. Cancer Epidemiology, Biomarkers & Prevention. doi: 10.1158/1055-9965.EPI-1-0084.
Hecht, S. S., Carmella, S. G., Murphy, S. E., Riley, W. T., Le, C., Luo, X., Mooney, M., & Hatuskami, D. K. (2007). Similar exposure to a tobacco-specific carcinogen in smokeless tobacco users and cigarette smokers. Cancer Epidemiology, Biomarkers & Prevention, 16, 1567-1572.
Hatsukami, D. K., Benowitz, N. L., Rennard, S. I., Oncken, C., & Hecht, S. S. (2006). Biomarkers to assess the utility of potential reduced exposure products. Nicotine Tobacco Research, 4, 600-622.
Bernert, J. T., Pirkle, J. L., Xia, Y., Jain, R. B., Ashley D. L., & Sampson, E. J. (2010). Urine concentrations of a tobacco-specific nitrosamine carcinogen in the U.S. population from secondhand smoke exposure. Cancer Epidemiology, Biomarkers & Prevention, 19(11), 2969-77
Xia, Y., Jain, R., Bernert, J. T., Ashley, D. L., & Pirkle, J. L. (2011). Tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) in smokers in the United States: NHANES 2007-2008. Biomarkers, 16(2), 112-119.
Protocol ID
720402
Variables
Export VariablesVariable Name | Variable ID | Variable Description | dbGaP Mapping |
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Measure Name
NNAL in Urine
Release Date
May 3, 2024
Definition
An assay to measure NNAL, the primary metabolite of NNK.
Purpose
To measure exposure to tobacco-specific nitrosamine carcinogens.
Keywords
biomarker, NNAL, urine, tobacco-specific carcinogen, TSNA, tobacco smoke exposure, tobacco product exposure, tobacco chemical exposure, smokeless tobacco exposure, laboratory protocol, second hand smoke, secondhand smoke, second-hand smoke
Measure Protocols
Protocol ID | Protocol Name |
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720402 | NNAL in Urine |
Publications
There are no publications listed for this protocol.