Lab Testing for Nicotine
Cotinine measurement is the gold standard lab test for nicotine testing, as it is the best biomarker of tobacco exposure with a long half-life and can be measured in blood, saliva, hair, and urine. 1
Primary Testing Method: Cotinine
Cotinine is regarded as the best biomarker of tobacco exposure because it has a half-life of 16-18 hours and can be biochemically verified for up to 7 days after use. 1 This nicotine metabolite provides superior sensitivity compared to other markers and can be measured in multiple specimen types. 1
Specimen Types and Cutoff Values
The American Thoracic Society provides specific cutoff values for cotinine testing across different specimens: 1
- Urine cotinine: 30 ng/mL cutoff for positive result
- Serum/plasma cotinine: 3-5 ng/mL cutoff for positive result
- Saliva cotinine: 3-5 ng/mL cutoff for positive result
Key Advantage and Limitation
Cotinine is more sensitive than carbon monoxide but has one important limitation: it cannot distinguish nicotine replacement therapy (NRT) from tobacco use, making it less specific in patients using nicotine patches, gum, or other cessation products. 1
Alternative Testing Methods
Carbon Monoxide (CO)
- Measured in expired breath with a cutoff of 5-6 ppm 1
- Half-life of 2-8 hours with a biochemically verifiable window of only 1 day 1
- Can be measured in breath up to 24-48 hours after smoking 1
- Reasonably sensitive for recent or heavy cigarette use but does not detect smokeless tobacco or NRT 1
- Important caveat: With the rise of smoking non-tobacco products (e.g., marijuana), a positive CO level may not be sufficient to conclude tobacco smoking; combining CO with urine drug screen and/or urine cotinine provides more robust assessment 1
NNAL (Tobacco-Specific Nitrosamine Metabolite)
- Measured in urine only with a cutoff of 47.3 pg/mL 1
- Half-life of 10-18 days and can be detected in urine for 6-12 weeks 1
- Highly tobacco-specific and can distinguish NRT from tobacco use 1
- Detects smokeless tobacco use 1
Laboratory Methods
Modern quantitative testing uses liquid chromatography-tandem mass spectrometry (LC-MS/MS) for simultaneous determination of nicotine and its metabolites (cotinine and 3-OH-cotinine) in urine. 2, 3 This method:
- Requires only 60 μL of human urine 2
- Provides analytical ranges of 10-2500 ng/mL for nicotine and cotinine 2
- Offers precision better than 10% for major nicotine analytes 3
- Allows analysis of over 170 samples per day 4
Clinical Application Considerations
When ordering nicotine testing, specify cotinine measurement in urine as the first-line test due to its optimal balance of sensitivity, detection window, and ease of collection. 1 For patients using NRT, consider adding NNAL testing to distinguish therapeutic nicotine use from tobacco exposure. 1
The National Health and Nutrition Examination Survey uses serum cotinine concentration as the standard biomarker for assessing tobacco smoke exposure in population studies. 1