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EPIC CODE: LAB3971 Nicotine and Metabolites, Random, Urine

Additional Codes

Sunquest:    NICOUM

Mayo:           NICOU

Reporting Name

Nicotine and Metabolites, U

Useful For

Monitoring tobacco use


Monitoring patients on nicotine-replacement therapy for concurrent use of tobacco products

Performing Laboratory

Mayo Clinic Laboratories in Rochester

Specimen Type


Specimen Required

Supplies: Sarstedt Aliquot Tube, 5 mL (T914)

Container/Tube: Plastic, 5 mL, aliquot tube

Specimen Volume: 3 mL

Collection Instructions:

1. Collect a random urine specimen.

2. No preservative.

Specimen Minimum Volume

2 mL

Specimen Stability Information

Specimen Type Temperature Time Special Container
Urine Refrigerated (preferred) 28 days
  Frozen  365 days
  Ambient  28 days

Reference Values

Non-tobacco user with no passive exposure:


<5.0 ng/mL



<5.0 ng/mL



<2.0 ng/mL



<2.0 ng/mL

Day(s) Performed

Monday through Thursday

Test Classification

This test was developed, and its performance characteristics determined by Mayo Clinic in a manner consistent with CLIA requirements. This test has not been cleared or approved by the US Food and Drug Administration.

CPT Code Information


G0480 (if appropriate)

LOINC Code Information

Test ID Test Order Name Order LOINC Value
NICOU Nicotine and Metabolites, U 43125-4


Result ID Test Result Name Result LOINC Value
82510 Nicotine 3854-7
21321 Cotinine 10366-3
21324 Nornicotine 33917-6
21323 Anabasine 33915-0

Clinical Information

Tobacco use is the leading cause of death in the United States. Nicotine, coadministered in tobacco products such as cigarettes, pipes, cigars, or chew, is an addictive substance that causes individuals to continue use of tobacco despite concerted efforts to quit. Nicotine stimulates dopamine release and increases dopamine concentration in the nucleus accumbens, a mechanism that is thought to be the basis for addiction to drugs of abuse.


Nicotine is rapidly metabolized in the liver to cotinine, exhibiting an elimination half-life of 2 hours. Cotinine exhibits an apparent elimination half-life of 15 hours. Patients using tobacco products excrete nicotine in urine in the concentration range of 1000 to 5000 ng/mL. Cotinine accumulates in urine in proportion to dose and hepatic metabolism (which is genetically determined); most tobacco users excrete cotinine in the range of 1000 to 8000 ng/mL. Urine concentrations of nicotine and metabolites in these ranges indicate the subject is using tobacco or is receiving high-dose nicotine patch therapy.


In addition to nicotine and metabolites, tobacco products contain other alkaloids that can serve as unique markers of tobacco use. Two such markers are anabasine and nornicotine. Anabasine is present in tobacco products but not in nicotine replacement therapies. Nornicotine is present as an alkaloid in tobacco products and as a metabolite of nicotine. The presence of anabasine (>10 ng/mL) or nornicotine (>30 ng/mL) in urine indicates current tobacco use, irrespective of whether the subject is on nicotine replacement therapy. The presence of nornicotine without anabasine is consistent with use of nicotine replacement products. Heavy tobacco users who abstain from tobacco for 2 weeks exhibit urine nicotine values below 30 ng/mL, cotinine values below 50 ng/mL, anabasine levels below 2 ng/mL, and nornicotine levels below 2 ng/mL.


Passive exposure to tobacco smoke can cause accumulation of nicotine metabolites in nontobacco users. Urine cotinine has been observed to accumulate up to 20 ng/mL from passive exposure. Neither anabasine nor nornicotine accumulates from passive exposure.


Tobacco users engaged in programs to abstain from tobacco require support in the form of counseling, pharmacotherapy, and continuous encouragement. Occasionally, counselors may elect to monitor abstinence by biochemical measurement of nicotine and metabolites in a random urine specimen to verify abstinence. If results of biologic testing indicate the patient is actively using a tobacco product during therapy, additional counseling or intervention may be appropriate.


Quantification of urine nicotine and metabolites, while a patient is actively using a tobacco product, is useful to define the concentrations that a patient achieves through self-administration of tobacco. Nicotine replacement dose can then be tailored to achieve the same concentrations early in treatment to assure adequate nicotine replacement so the patient may avoid the strong craving they may experience early in the withdrawal phase. This can be confirmed by measurement of urine nicotine and metabolite concentrations at a steady state (2-3 days after replacement therapy is started). Once the patient is stabilized on the dose necessary to achieve complete replacement and responding well to therapy, the replacement dose can be slowly tapered to achieve complete withdrawal.


Urine nicotine in the range of 1000 to 5000 ng/mL with cotinine in the range of 1000 to 8000 ng/mL indicates the subject is either actively using a tobacco product or on high-dose nicotine patch therapy.


The presence of anabasine and nornicotine indicates a subject on patch therapy who is actively using a tobacco product.


Typical findings are as follows:


While using a tobacco product:

-Peak nicotine concentration: 1000 to 5000 ng/mL

-Peak cotinine concentration: 1000 to 8000 ng/mL

-Anabasine concentration: 10 to 500 ng/mL

-Nornicotine concentration: 30 to 900 ng/mL


Tobacco user after 2 weeks complete abstinence:

-Nicotine concentration: <30 ng/mL

-Cotinine concentration: <50 ng/mL

-Anabasine concentration: <2.0 ng/mL

-Nornicotine concentration: <2.0 ng/mL


Nontobacco user with passive exposure:

-Nicotine concentration: <20 ng/mL

-Cotinine concentration: <20 ng/mL

-Anabasine concentration: <2.0 ng/mL

-Nornicotine concentration: <2.0 ng/mL


Nontobacco user with no passive exposure:

-Nicotine concentration: <5.0 ng/mL

-Cotinine concentration: <5.0 ng/mL

-Anabasine concentration: <2.0 ng/mL

-Nornicotine concentration: <2.0 ng/mL


Knowledge of time elapsed between last dose and specimen collection is important for interpretation of test results.

Clinical Reference

1. Dale LC, Hurt RD, Hays JT: Drug therapy to aid in smoking cessation. Tips on maximizing patients' chances for success. Postgrad Med. 1998 Dec;104(6):75-78, 83-84

2. Rudasingwa G, Kim Y, Lee C, Lee J, Kim S, Kim S. Comparison of nicotine dependence and biomarker levels among traditional cigarette, heat-not-burn cigarette, and liquid e-cigarette users: Results from the Think Study. Int J Environ Res Public Health. 2021 Apr 29;18(9):4777. doi: 10.3390/ijerph18094777

3. Sharma P, Sane N, Anand SD, Marimutthu P, Benegal V. Assessment of cotinine in urine and saliva of smokers, passive smokers, and nonsmokers: Method validation using liquid chromatography and mass spectrometry. Indian J Psychiatry. 2019 May-Jun;61(3):270-276. doi: 10.4103/psychiatry.IndianJPsychiatry_61_18

Method Description

Nicotine and metabolites are extracted from urine by solid-phase extraction techniques. The extract eluate is quantified by high-performance liquid chromatography-tandem mass spectrometry.(Moyer TP, Charlson JR, Enger RJ, et al: Simultaneous analysis of nicotine, nicotine metabolites, and tobacco alkaloids in serum or urine by tandem mass spectrometry, with clinically relevant metabolic profiles. Clin Chem. 2002 Sep;48(9):1460-1471; Oh J, Park MS, Chun MR, et al: A simple and high-throughput LC-MS/MS method for simultaneous measurement of nicotine, cotinine, 3-OH cotinine, nornicotine, and anabasine in urine and its application in the general Korean population. J Anal Toxicol. 2022 Feb 14:46(1):25-36. doi: 10.1093/jat/bkaa177

Report Available

2 to 5 days

Specimen Retention Time

2 weeks

Reject Due To

Gross hemolysis OK
Gross icterus OK

NY State Approved


Method Name

Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS)


If not ordering electronically, complete, print, and send a Therapeutics Test Request (T831) with the specimen.