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Cited by 3Hormonal and reproductive studiesSmoking Behavior and CessationSperm and Testicular Function

Association between serum cotinine and total testosterone in adult males based on NHANES 2011–2016

Open access
Liu, Ming, Gou, Yunpeng, Zou, Bing, Li, Xianhui, Yang, Ping·2024·Scientific Reports
DOI 10.1038/s41598-024-74482-6PMC PMC11450214
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Abstract

The relationship between smoking and testosterone levels in adult males remains a topic of ongoing debate. Serum cotinine is considered a reliable marker of both smoking intensity and exposure to tobacco smoke. Therefore, we aim to examine the association between serum cotinine levels and total testosterone concentrations in adult males using data from the U.S. National Health and Nutrition Examination Survey (NHANES) database. Our study assessed the relationship between serum cotinine and total testosterone using weighted linear regression models and subgroup analysis. A fully adjusted model with smooth curve fitting was employed to investigate the potential nonlinear association between serum cotinine and total testosterone. Threshold effects were analyzed to identify the inflection point between serum cotinine and total testosterone. Indeed, a total of 7797 participants were included in our study. After adjusting for potential confounding variables, the findings indicate a positive association between serum cotinine levels and total testosterone levels (β: 0.05, 95%CI: 0.02, 0.09). Furthermore, applying smoothed curve fitting analysis and threshold effects, an inflection point was detected at a serum cotinine level of 487 ng/ml. Above this threshold, total testosterone levels declined with increasing serum cotinine levels. In conclusion, the findings of our study suggest a positive association between elevated serum cotinine levels and total testosterone levels in adult men. However, it is essential to note that this association may be reversed at excessively high serum cotinine concentrations.

AI snapshot

Written from the paper’s full text.

TL;DR

In adult males from NHANES 2011–2016, higher serum cotinine was associated with higher total testosterone after adjustment. However, the association reversed above an inflection point of 487 ng/mL serum cotinine.

Key findings

  • Among 7,797 adult males, fully adjusted weighted linear regression showed total testosterone increased by β 0.05 (95% CI 0.02, 0.09) per unit increase in serum cotinine.
  • Compared with the lowest cotinine quartile (Q1 ≤ 0.011 ng/mL), Q4 (> 57.7 ng/mL) had 35.94 ng/dL higher total testosterone (95% CI 22.71, 49.16; p for trend < 0.05).
  • Smooth curve fitting found a nonlinear relationship with an inflection point at serum cotinine 487 ng/mL: below 487 ng/mL, β 0.06 (95% CI −0.01, 0.12), p = 0.089; at or above 487 ng/mL, β −0.28 (95% CI −0.49, −0.08), p = 0.007.
  • Subgroup interaction tests were significant for age and race (p for interaction < 0.05); the positive association was stronger in ages 18–30 than 30–50 (adjusted β 0.08 vs. 0.07) and among Non-Hispanic Asians.

Methods

  • Cross-sectional analysis of NHANES 2011–2016; after excluding females, participants under 18, and those missing serum cotinine or total testosterone, the final sample was 7,797 adult males.
  • Serum cotinine was measured by ID HPLC-APCI MS/MS; total testosterone was measured by ID-LC-MS/MS.
  • Weighted univariate and multiple linear regression models were used, including unadjusted, minimally adjusted (age and race), and fully adjusted models (age, race, education, marital status, alcohol consumption, hypertension, diabetes, physical activity, and BMI), plus subgroup analyses, smooth curve fitting, and threshold-effect analysis.

Limitations

  • Stated: the study examined only total testosterone, not free testosterone or SHBG; it is unclear whether the association is reversible after smoking cessation; the cross-sectional design limits causal inference; unidentified confounders may remain; results may not generalize to individuals under 18; and the analysis relied on secondary data.
  • Evident from the design: cotinine and testosterone were measured at a single time point, and smoking status, alcohol use, hypertension, diabetes, and physical activity relied partly on self-report or medication use.

Model-written summary. Check it against the paper before relying on it.

Integrity signals

Registry

Crossref · 10.1038/s41598-024-74482-6

Cites

41 of 41 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Claims the paper makes

8 of 20 cited sources checked · 4 supported · 0 flagged · accuracy not yet measured

  • Premisecitation

    Serum cotinine is directly proportional to the extent of tobacco smoke exposure.

    “Serum cotinine, a primary metabolite of nicotine, is directly proportional to the extent of tobacco smoke exposure and can serve as a reliable indicator of smoking intensity and tobacco smoke exposure18,19.”
    Quote in the paperSubstantial claim1 of 2 sources checked
    • cites [18] Florescu, A. et al. Methods for quantification of exposure to cigarette smoking and environmental tobacco smoke: focus on developmental toxicology. Ther. Drug Monit.31, 14–30. https://doi.org/10.1097/FTD.0b013e3181957a3b (2009). source

      The Biomarkers section (page not provided) says blood cotinine approximates absorbed nicotine dose and that cotinine levels reflect tobacco-smoke exposure. It also describes variation in the relationship between cotinine levels and nicotine intake, so it does not establish that serum cotinine is directly proportional to the extent of exposure. (LLM, conf=0.86)

      Experimental — accuracy not yet measured
    • cites [19] Schick, S. F. et al. Biomarkers of exposure to new and emerging tobacco delivery products. Am. J. Physiol. Lung Cell. Mol. Physiol.313, L425–l452. https://doi.org/10.1152/ajplung.00343.2016 (2017). source
  • Premisecitation

    Serum cotinine can serve as a reliable indicator of smoking intensity and tobacco smoke exposure.

    “Serum cotinine, a primary metabolite of nicotine, is directly proportional to the extent of tobacco smoke exposure and can serve as a reliable indicator of smoking intensity and tobacco smoke exposure18,19.”
    • Also stated in: § Abstract · plain
      “Serum cotinine is considered a reliable marker of both smoking intensity and exposure to tobacco smoke.”
    Quote in the paperSubstantial claim1 of 2 sources checked
    • cites [18] Florescu, A. et al. Methods for quantification of exposure to cigarette smoking and environmental tobacco smoke: focus on developmental toxicology. Ther. Drug Monit.31, 14–30. https://doi.org/10.1097/FTD.0b013e3181957a3b (2009). source
    • cites [19] Schick, S. F. et al. Biomarkers of exposure to new and emerging tobacco delivery products. Am. J. Physiol. Lung Cell. Mol. Physiol.313, L425–l452. https://doi.org/10.1152/ajplung.00343.2016 (2017). source
  • Findingown data

    In people aged 50 and above, there is an inflection point at serum cotinine = 508 ng/ml.

    “After stratified analysis by age, we also found an inflection point of 508 in people aged 50 and above, as shown in Table 3.”
    Quote in the paperSubstantial claim
  • Findingown data

    In adult males, after adjusting for age, race, education, marital status, alcohol consumption, hypertension, diabetes, physical activity status, and BMI, serum cotinine levels are positively associated with total testosterone levels (β 0.05, 95%CI 0.02, 0.09).

    “After adjusting for potential confounding variables, the findings indicate a positive association between serum cotinine levels and total testosterone levels (β: 0.05, 95%CI: 0.02, 0.09).”
    • Also stated in: § Association between serum cotinine and total testosterone · plain
      “In the model 3, the fully adjusted model that adjusts for age, race, education, marital status, alcohol consumption, hypertension, diabetes, physical activity status, and BMI showed that the total testosterone was increased by 0.05 (0.02, 0.09) for each additional unit of serum cotinine with.”
    • Also stated in: § Discussion · boosted
      “The research demonstrates a positive association between elevated serum cotinine levels and total testosterone levels in adult males. This association remained statistically significant following adjustments for potential confounding variables.”
    Quote in the paperSubstantial claim
  • Findingown data

    In the 30-50 age group, the relationship between serum cotinine and testosterone was a linear positive association.

    “Conversely, in the 18–30 and 30–50 age groups, the relationship between serum cotinine and testosterone was a linear positive association.”
    Quote in the paperSubstantial claim
  • Classified as probably not a substantial claim. 7
  • May combine two claims. 5
  • The quote only partly states this claim. 2

  • Classified as probably not a substantial claim. May combine two claims.

    Premisecitation

    Serum testosterone levels decline with age, commencing in men in their mid-30s and decreasing at an average annual rate of 1.6%.

    “We speculate that this phenomenon may be attributed to the gradual decline in serum testosterone levels with age, commencing in men in their mid-30s and decreasing at an average annual rate of 1.6%38–43.”
    Quote in the paperSubstantial claim2 of 6 sources checked
    • cites [38] Tajar, A. et al. Characteristics of secondary, primary, and compensated hypogonadism in aging men: evidence from the European Male Ageing Study. J. Clin. Endocrinol. Metab.95, 1810–1818. https://doi.org/10.1210/jc.2009-1796 (2010). source
    • cites [39] Feldman, H. A. et al. Age trends in the level of serum testosterone and other hormones in middle-aged men: longitudinal results from the Massachusetts male aging study. J. Clin. Endocrinol. Metab.87, 589–598. https://doi.org/10.1210/jcem.87.2.8201 (2002). source
    • cites [40] Nieschlag, E. et al. Investigation, treatment and monitoring of late-onset hypogonadism in males: ISA, ISSAM, and EAU recommendations. Int. J. Androl.28, 125–127. https://doi.org/10.1111/j.1365-2605.2005.00553.x (2005). source
    • cites [41] Harman, S. M., Metter, E. J., Tobin, J. D., Pearson, J. & Blackman, M. R. Longitudinal effects of aging on serum total and free testosterone levels in healthy men. Baltimore Longitudinal Study of Aging. J. Clin. Endocrinol. Metab.86, 724–731. https://doi.org/10.1210/jcem.86.2.7219 (2001). source

      The Discussion reports that serum testosterone declined progressively with age in the men studied, from the third through ninth decades. It does not identify the mid-30s as the onset or report an average decline of 1.6% per year; it gives a total-testosterone decline of 0.110 nmol/L (3.2 ng/dL) per year instead. Page numbers were not available in our copy. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
    • cites [42] Stanworth, R. D. & Jones, T. H. Testosterone for the aging male; current evidence and recommended practice. Clin. Interv Aging. 3, 25–44. https://doi.org/10.2147/cia.s190 (2008). source

      The “Changes in testosterone levels with aging” section reports that total testosterone declines by an average of 1.6% per year. It does not say that the decline begins in men in their mid-30s; that onset age is unsupported by this source. The citing authors’ proposed explanation of their phenomenon is their own speculation, not a finding attributed to this paper. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
    • cites [43] Huhtaniemi, I. & Forti, G. Male late-onset hypogonadism: pathogenesis, diagnosis and treatment. Nat. Rev. Urol.8, 335–344. https://doi.org/10.1038/nrurol.2011.47 (2011). source

  • Classified as probably not a substantial claim.

    Premisecitation

    Numerous studies have examined the impact of smoking on male reproductive hormone levels, yielding conflicting results.

    “Numerous studies have examined the impact of smoking on male reproductive hormone levels, yielding conflicting results.”
    Quote in the paperSubstantial claim2 of 6 sources checked
    • cites [8] Dai, J. B., Wang, Z. X. & Qiao, Z. D. The hazardous effects of tobacco smoking on male fertility. Asian J. Androl.17, 954–960. https://doi.org/10.4103/1008-682x.150847 (2015). source
    • cites [12] Halmenschlager, G., Rossetto, S., Lara, G. M. & Rhoden, E. L. Evaluation of the effects of cigarette smoking on testosterone levels in adult men. J. Sex. Med.6, 1763–1772. https://doi.org/10.1111/j.1743-6109.2009.01227.x (2009). source
    • cites [14] Schooling, C. M. Smoking, sex, risk factors and abdominal aortic aneurysm: is it all down to testosterone? J. Epidemiol. Community Health. 69, 495. https://doi.org/10.1136/jech-2015-205595 (2015). source
    • cites [15] Wu, F. C. et al. Hypothalamic-pituitary-testicular axis disruptions in older men are differentially linked to age and modifiable risk factors: the European Male Aging Study. J. Clin. Endocrinol. Metab.93, 2737–2745. https://doi.org/10.1210/jc.2007-1972 (2008). source
    • cites [16] Zhao, J., Leung, J. Y. Y., Lin, S. L. & Mary Schooling, C. Cigarette smoking and testosterone in men and women: a systematic review and meta-analysis of observational studies. Prev. Med.85, 1–10. https://doi.org/10.1016/j.ypmed.2015.12.021 (2016). source
    • cites [17] Holmboe, S. A. et al. Use of e-cigarettes associated with lower sperm counts in a cross-sectional study of young men from the general population. Hum. Reprod.35, 1693–1701. https://doi.org/10.1093/humrep/deaa089 (2020). source

      The Introduction (page unavailable) cites several studies of cigarette smoking and male hormone levels, supporting that the topic has been studied. It does not characterize their hormone findings as conflicting; instead, it says smokers are well known to have higher testosterone levels, and the Discussion says this study’s findings accord with other population-based studies. The Introduction’s reference to conflicting results concerns marijuana and semen quality, not smoking and hormones. (LLM, conf=0.84)

      Experimental — accuracy not yet measured

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    Smoking can have adverse effects on male reproductive health, including reduced sperm count and diminished semen quality.

    “It is widely acknowledged that smoking can have adverse effects on male reproductive health, including reduced sperm count and diminished semen quality8–11.”
    Quote in the paperNot a substantial claim2 of 4 sources checked
    • cites [8] Dai, J. B., Wang, Z. X. & Qiao, Z. D. The hazardous effects of tobacco smoking on male fertility. Asian J. Androl.17, 954–960. https://doi.org/10.4103/1008-682x.150847 (2015). source
    • cites [9] Sansone, A. et al. Smoke, alcohol and drug addiction and male fertility. Reprod. Biol. Endocrinol.16https://doi.org/10.1186/s12958-018-0320-7 (2018). source
    • cites [10] Teles, A. G. et al. Prevalence, severity, and risk factors for erectile dysfunction in a representative sample of 3,548 Portuguese men aged 40 to 69 years attending primary healthcare centers: results of the Portuguese erectile dysfunction study. J. Sex. Med.5, 1317–1324. https://doi.org/10.1111/j.1743-6109.2007.00745.x (2008). source
    • cites [11] Ramlau-Hansen, C. H. et al. Is smoking a risk factor for decreased semen quality? A cross-sectional analysis. Hum. Reprod.22, 188–196. https://doi.org/10.1093/humrep/del364 (2007). source

  • May combine two claims.

    Findingown data

    The interaction tests for marital status, BMI, and alcohol consumption were not significant (p for interaction >0.05).

    “However, the interaction tests for marital status, BMI, and alcohol consumption were not significant (p for interaction > 0.05).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingown data

    The positive association was stronger in the 18-30 age group, where testosterone levels increased consistently with rising serum cotinine levels.

    “This positive association was stronger in the 18–30 age group, where testosterone levels increased consistently with rising serum cotinine levels.”
    Quote in the paperSubstantial claim

  • May combine two claims. The quote only partly states this claim.

    Findingown data

    In the fully adjusted model, participants in the highest serum cotinine quartile (Q4, >57.7 ng/ml) had 35.94 ng/dL higher total testosterone than those in the lowest quartile (Q1, ≤0.011 ng/ml) (95%CI 22.71,49.16), with p for trend <0.05.

    “participants in Q4 had 35.94 ng/dL higher total testosterone than Q1(p for trend < 0.05).”
    Quote in the paperSubstantial claim

  • The quote only partly states this claim.

    Findingown data

    In people aged 50 and above, when serum cotinine is ≥508 ng/ml, total testosterone decreases as serum cotinine increases (β -0.37, 95%CI -0.72,-0.03, p=0.034).

    “in the group aged 50 and above, the association between serum cotinine and testosterone demonstrated a significant nonlinear relationship, where testosterone levels decreased as serum cotinine increased when serum cotinine concentration exceeded 508 ng/ml.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Interpretationreasoning

    This study is cross-sectional, limiting the ability to establish causality.

    “Furthermore, it is essential to note that this study is cross-sectional, limiting the ability to establish causality.”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim.

    Study design and conductnone given

    An appropriate examination weight was applied to represent the complex survey design.

    “An appropriate examination weight was applied to represent the complex survey design.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Interpretationreasoning

    The results may not be generalizable to individuals under the age of 18 due to the age restrictions of the NHANES 2011-2016 questionnaire.

    “It is also worth mentioning that the results may not be generalizable to individuals under the age of 18 due to the age restrictions of the NHANES 2011–2016 questionnaire.”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim.

    Study design and conductown data

    The fully adjusted model adjusted for age, race, education, marital status, alcohol consumption, hypertension, diabetes, physical activity status, and BMI.

    “In the model 3, the fully adjusted model that adjusts for age, race, education, marital status, alcohol consumption, hypertension, diabetes, physical activity status, and BMI showed that the total testosterone was increased by 0.05 (0.02, 0.09) for each additional unit of serum cotinine with.”
    Quote in the paperSubstantial claim
11 of 11

  • The paper relies on this without support.

    NHANES 2011–2016 is a nationally representative survey of the US population.

    “NHANES is a cross-sectional survey nationally representative of nutrition and health conditions in the United States, administered by the National Centers for Disease Control (CDC) and the National Center for Health Statistics of Prevention.”
1 of 1

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

Each contradiction between two passages listed here was confirmed by an independent check.

  • The 30-50 age group is described as having a positive association, but both reported slopes are negative

    Major

    The discussion calls the association in the 30-50 age group linear and positive. Table 3 reports negative estimated slopes on both sides of that age group's inflection point, although neither slope is statistically significant.

1 of 1

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Related

Authors
Liu, M., Gou, Y., Zou, B., Li, X., & Yang, P.
Corresponding author
Yang, Ping
ORCID
Venue
Scientific Reports
Publisher
Springer Science and Business Media LLC
Published
2024-10-04
Volume
14
Issue
1
Pages
23042-23042
ISSN
2045-2322 (ISSN-L)
DOI
10.1038/s41598-024-74482-6
PMC
PMC11450214
OpenAlex
W4403094021
Cited by
3
References
41
Citations by year
2025: 3
FWCI
0.88
Type
journal article
Language
en
Open access
Licence
Locations
  • Scientific Reports (publishedVersion, cc-by-nc-nd)
  • PubMed Central (publishedVersion, cc-by-nc-nd)
  • PubMed (publishedVersion)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
Topics
  • Health Sciences › Medicine › Endocrinology, Diabetes and Metabolism › Hormonal and reproductive studies
  • Health Sciences › Medicine › Physiology › Smoking Behavior and Cessation
  • Health Sciences › Medicine › Reproductive Medicine › Sperm and Testicular Function
Keywords
serum cotinine, total testosterone, smoking, tobacco smoke exposure, adult male, nonlinear relationship, threshold effects, inflection point, NHANES
MeSH terms
Adult, Aged, Cotinine, Humans, Male, Middle Aged, Nutrition Surveys, Smoking, Testosterone, United States, Biomarkers, Young Adult

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Classified by jev-1.13-20260917 (gold-guideline-v0.2) on 2026-11-05.

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