Analysis of urinary tobacco-specific nitrosamine 4- (methylnitrosamino)1-(3-pyridyl)-1- butanol (NNAL) and HPV infection in American women: National health and nutrition examination survey.

Tobacco-specific nitrosamines (TSNAs) are a group of toxic substances specific to tobacco. 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) is a tobacco-specific nitrosamine measurable in urine with a much longer half-life than cotinine. We aimed to examine the association between urinary tobacco-specific NNAL and HPV infection among American women. We used cross-sectional data from the National Health and Nutrition Examination Survey (NHANES) between 2007 and 2014 to collect details on their urinary NNAL, HPV infection status, and other essential variables. The association between dietary urinary NNAL and HPV infection status was analyzed by using a weighted multivariate logistic regression model, and stratified subgroup analysis. In total, 5197 participants aged 18-59 years were identified, with overall prevalence of high-risk and low-risk HPV infection of 22.0% and 19.1%, respectively. The highest quartile of NNAL(Q4) was more positively associated with low-risk HPV infection than the lowest quartile of NNAL(Q1) (OR = 1.83 (1.35,2.50), p<0.001). the highest quartile of NNAL(Q4) was more positively associated with high-risk HPV infection than the lowest quartile of NNAL(Q1) (OR = 2.20 (1.57,3.08), p < 0.001). In subgroup analyses, the positive correlation between urinary NNAL levels and low-risk HPV infection status was inconsistent in marital status and BMI (interaction p < 0.05). The positive association of urinary NNAL levels with high-risk HPV infection status was inconsistent in smoking and BMI. (interaction p < 0.05). Tobacco-specific NNAL levels positively correlate with high- and low-risk HPV. Future well-designed longitudinal studies are still needed to validate the effect of tobacco exposure on HPV infection by NNAL.

Comparison of bladder carcinogenesis biomarkers in the urine of traditional cigarette users and e-cigarette users.

Background: During the use of electronic cigarettes (e-cigarettes), users are still exposed to carcinogens similar to those found in tobacco products. Since these carcinogens are metabolized and excreted in urine, they may have carcinogenic effects on the bladder urinary tract epithelium. This meta-analysis aimed to compare bladder cancer carcinogens in the urine of tobacco users and e-cigarette users using a large number of samples. Methods: A systematic meta-analysis was performed using data obtained from several scientific databases (up to November 2023). This cumulative analysis was performed following the Preferred Reporting Items for Systematic Evaluation and Meta-Analysis (PRISMA) and Assessing the Methodological Quality of Systematic Evaluations (AMSTAR) guidelines, according to a protocol registered with PROSPERO. This study was registered on PROSPERO and obtained the unique number: CRD42023455600. Results: The analysis included 10 high-quality studies that considered polycyclic aromatic hydrocarbons (PAHs), volatile organic compounds (VOCs) and tobacco-specific nitrosamines (TSNAs). Statistical indicators show that there is a difference between the tobacco user group and the e-cigarette user group in terms of 1-Hydroxynaphthalene (1-NAP) [weighted mean difference (WMD)10.14, 95% confidence interval (CI) (8.41 to 11.88), p < 0.05], 1-Hydroxyphenanthrene (1-PHE) [WMD 0.08, 95% CI (-0.14 to 0.31), p > 0.05], 1-Hydroxypyrene (1-PYR) [WMD 0.16, 95% CI (0.12 to 0.20), p < 0.05], 2-Hydroxyfluorene (2-FLU) [WMD 0.69, 95% CI (0.58 to 0.80), p < 0.05], 2-Hydroxynaphthalene (2-NAP) [WMD 7.48, 95% CI (4.15 to 10.80), p < 0.05], 3-Hydroxyfluorene (3-FLU) [WMD 0.57, 95% CI (0.48 to 0.66), p < 0.05], 2-Carbamoylethylmercapturic acid (AAMA) [WMD 66.47, 95% CI (27.49 to 105.46), p < 0.05], 4-Hydroxy-2-buten-1-yl-mercapturic acid (MHBMA) [WMD 287.79, 95% CI (-54.47 to 630.04), p > 0.05], 4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNAL) [WMD 189.37, 95% CI (78.45 to 300.29), p < 0.05], or N0-nitrosonornicotine (NNN) [WMD 11.66, 95% CI (7.32 to 16.00), p < 0.05]. Conclusion: Urinary bladder cancer markers were significantly higher in traditional tobacco users than in e-cigarette users.Systematic review registration: PROSPERO (CRD42023455600: https://www.crd.york.ac.uk/PROSPERO/).

Biomarkers of Toxic Exposure and Oxidative Stress Among U.S. Adult Users of Premium Cigar Versus Other Cigar Subtypes: 2013-2019.

INTRODUCTION: Cigars are currently the second-highest-used combustible tobacco product among U.S. adults, but knowledge about health effects of premium cigars versus other cigar subtype use is limited. AIMS AND METHODS: This study analyzed the biospecimen data (n = 31 875) from Waves 1-5 of the Population Assessment of Tobacco and Health Study, collected during 2013-2019. Multivariable generalized estimation equations, accounting for within-person clustering, were conducted to examine differences in urine biomarkers of exposure (BOE) from five classes of harmful and potentially harmful constituents along with a biomarker of oxidative stress (urine 8-isoprostane) among exclusive users of premium cigars versus other exclusive cigar subtypes (ie, non-premium large cigars, cigarillos, and filtered cigars), cigarettes, and non-tobacco users. RESULTS: In comparison to non-tobacco users, exclusive premium cigar users had higher geometric mean concentrations of the nicotine metabolite cotinine (5.8 vs. 0.5ng/mg, p < .0001), tobacco-specific nitrosamine (TSNA) (4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL): 7.8 vs. 1.3pg/mg, p < .0001), and volatile organic compound (VOC) (N-Acetyl-S-(2-cyanoethyl)-L-cysteine (CYMA, acrylonitrile): 4.7 vs. 1.6ng/mg, p < .0001). Exclusive premium cigar users were less likely to be daily users than other tobacco user groups and had comparable BOEs with exclusive non-premium large cigar users but generally lower BOEs than exclusive cigarillo, filtered cigar, and cigarette smokers. Daily exclusive premium cigar users had similar nicotine and TSNA exposure but lower exposure to polycyclic aromatic hydrocarbons and volatile organic compounds than exclusive cigarillo and filtered cigar users. CONCLUSIONS: Premium cigar use exhibits different exposure to toxicants from other cigar subtype users. Regulations of premium cigars need to formalize product definition and take the population's health effects into consideration. IMPLICATIONS: This population study provides important information on BOE and potential harm with premium cigar use and its potential health effects. At present, premium cigars appear to pose a relatively low overall population health risk due to low frequency of use. However, future regulation of other tobacco products might change the landscape of premium cigar use and alter the overall health impact.

Comparison of a Tobacco-Specific Carcinogen in Tobacco Cigarette, Electronic Cigarette, and Dual Users.

BACKGROUND: 4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) is known as a lung carcinogen. The objective of this study was to investigate associations of urine NNAL concentrations and smoking status. METHODS: This was a cross-sectionally designed study based on data from the 2016-2018 Korean National Health and Nutrition Examination Survey. A total of 2,845 participants were classified into past-smoker, electronic cigarette (e-cigar) only, dual-user, and cigarette only smoker groups. All sampling and weight variables were stratified and analysis was conducted accounting for the complex sampling design. Analysis of covariance was used to compare the geometric mean of urine NNAL concentrations and log-transformed urine NNAL level among smoking status with weighted survey design. Post hoc paired comparisons with Bonferroni adjustment was performed according to smoking status. RESULTS: The estimated geometric mean concentrations of urine NNAL were 1.974 ± 0.091, 14.349 ± 5.218, 89.002 ± 11.444, and 117.597 ± 5.459 pg/mL in past-smoker, e-cigar only, dual-user, and cigarette only smoker groups, respectively. After fully adjusting, log-transformed urine NNAL level was significantly different among groups (P < 0.001). Compared with the past-smoker group, e-cigar only, dual-user, and cigarette only smoker groups showed significantly higher log-transformed urine NNAL concentrations in post hoc test (all P < 0.05). CONCLUSION: E-cigar only, dual-user, and cigarette only smoker groups showed significantly higher geometric mean concentrations of urine NNAL than the past-smoker group. Conventional cigarette, dual users, and e-cigar users can potentially show harmful health effects from NNAL.

Respiratory Exposure to Thirdhand Cigarette Smoke Increases Concentrations of Urinary Metabolites of Nicotine.

INTRODUCTION: The aims of this study were to characterize particle size in a thirdhand smoke (THS) aerosol and measure the effects of controlled inhalational exposure to THS on biomarkers of tobacco smoke exposure, inflammation, and oxidative stress in human subjects. Secondhand cigarette smoke changes physically and chemically after release into the environment. Some of the resulting chemicals persist indoors as thirdhand cigarette smoke. THS that is sorbed to surfaces can emit particles back into the air. AIMS AND METHODS: Smoke particle size was measured with a scanning mobility particle sizer and condensation particle counter. Using a crossover study design, 18 healthy nonsmokers received a 3-hour inhalational exposure to THS and to filtered, conditioned air. THS was generated with a smoking machine and aged overnight in a chamber. The chamber was flushed with clean air to create the THS aerosol. The tobacco smoke metabolites cotinine, 3-hydroxycotinine and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) were measured in urine. Vascular endothelial growth factor and interleukin-6 in plasma, and 8-isoprostane in urine, were measured using enzyme-linked immunosorbent assay kits. RESULTS: Mean smoke particle size increased with aging (171 to 265 nm). We found significant increases in urinary cotinine and 3-hydroxycotinine after 3 hours of exposure to THS and no significant increases in NNAL, interleukin-6, vascular endothelial growth factor or 8-isoprostane. CONCLUSIONS: Acute inhalational exposure to 22-hour old tobacco smoke aerosol caused increases in the metabolites of nicotine but not the metabolites of the tobacco-specific nitrosamine NNK (4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone). This corroborates the utility of cotinine and NNAL for secondhand and THS exposure screening. IMPLICATIONS: This study shows that a 3-hour inhalational exposure to the tobacco smoke aerosol that forms in a room that has been smoked in and left unventilated overnight causes increases in urinary metabolites of nicotine, but not of the tobacco-specific nitrosamine NNK. This suggests that cleaning personnel and others who live and work in rooms polluted with aged or thirdhand cigarette smoke regularly may have inhalational exposures and potential health effects related to their exposure to nicotine and other smoke toxicants.

Carcinogenic and tobacco smoke-derived particulate matter biomarker uptake and associated healthcare patterns among children.

BACKGROUND: The objective was to assess the associations of child tobacco smoke exposure (TSE) biomarkers (urinary cotinine, NNAL, and nicotelline N-oxides) and parent-reported smoking and child TSE patterns with total hospital visits, pediatric emergency department (PED) visits, urgent care (UC), revisits, and hospital admissions among 0-9-year-olds. METHODS: A convenience sample of PED/UC patients (N = 242) who presented to a large, US children's hospital who had baseline urine samples assayed for the TSE biomarkers of interest were included. Biomarker levels were log-transformed, and linear and Poisson regression models were built. RESULTS: The geometric means of child cotinine, creatinine-adjusted NNAL, and N-oxide levels were 11.2 ng/ml, 30.9 pg/mg creatinine, and 24.1 pg/ml, respectively. The mean (SD) number of daily cigarettes smoked by parents was 10.2 (6.1) cigarettes. Each one-unit increase in log-NNAL levels was associated with an increase in total UC visits (aRR = 1.68, 95% CI = 1.18-2.39) among 0-9-year-olds, while controlling for the covariates. Each one-unit increase in child log-NNAL/cotinine ratio (×103) values was associated with an increase in total hospital visits (aRR = 1.39, 95% CI = 1.10-1.75) and UC visits (aRR = 1.56, 95% CI = 1.14-2.13) over 6 months. CONCLUSION: Systematic screening for child TSE should be conducted during all hospital visits. The comprehensive assessment of TSE biomarkers should be considered to objectively measure young children's exposure. IMPACT: Higher levels of cotinine, a widely used tobacco smoke exposure biomarker, have been associated with higher healthcare utilization patterns among children. Less is known on the associations of carcinogenic and tobacco smoke-derived particulate matter biomarker uptake with child healthcare utilization patterns. This study assessed the associations of several biomarkers with healthcare utilization patterns among pediatric emergency department patients ages 0-9 years who lived with tobacco smokers. Higher urinary NNAL biomarker levels, in individual and ratio form with cotinine, increased children's risk for urgent care visits over 6 months. Higher parent-reported cumulative child tobacco smoke exposure increased children's risk for hospital admissions.

Association of Urinary Biomarkers of Smoking-Related Toxicants with Lung Cancer Incidence in Smokers: The Multiethnic Cohort Study.

BACKGROUND: While cigarette smoking is the leading cause of lung cancer, the majority of smokers do not develop the disease over their lifetime. The inter-individual differences in risk among smokers may in part be due to variations in exposure to smoking-related toxicants. METHODS: Using data from a subcohort of 2,309 current smokers at the time of urine collection from the Multiethnic Cohort Study, we prospectively evaluated the association of ten urinary biomarkers of smoking-related toxicants [total nicotine equivalents (TNE), a ratio of total trans-3'-hydroxycotinine (3-HCOT)/cotinine (a phenotypic measure of CYP2A6 enzymatic activity), 4-(methylnitrosamino)-1-3-(pyridyl)-1-butanol (NNAL), S-phenylmercapturic acid (SPMA), 3-hydroxypropyl mercapturic acid (3-HPMA), phenanthrene tetraol (PheT), 3-hydroxyphenanthrene (PheOH), the ratio of PheT/PheOH, cadmium (Cd), and (Z)-7-(1R,2R,3R,5S)-3,5-dihydroxy-2-[(E,3S)-3-hydroxyoct-1-enyl]cyclopenyl]hept-5-enoic acid (8-iso-PGF2α)] with lung cancer risk (n = 140 incident lung cancer cases over an average of 13.4 years of follow-up). Lung cancer risk was estimated using Cox proportional hazards models. RESULTS: After adjusting for decade of birth, sex, race/ethnicity, body mass index, self-reported pack-years, creatinine, and urinary TNE (a biomarker of internal smoking dose), a one SD increase in log total 3-HCOT/cotinine (HR, 1.33; 95% CI, 1.06-1.66), 3-HPMA (HR, 1.41; 95% CI, 1.07-1.85), and Cd (HR, 1.45; 95% CI, 1.18-1.79) were each associated with increased lung cancer risk. CONCLUSIONS: Our study demonstrates that urinary total 3-HCOT/cotinine, 3-HPMA, and Cd are positively associated with lung cancer risk. These findings warrant replication and consideration as potential biomarkers for smoking-related lung cancer risk. IMPACT: These biomarkers may provide additional information on lung cancer risk that is not captured by self-reported smoking history or TNE. See related commentary by Etemadi et al., p. 289.

Associations between urinary biomarkers of oxidative stress and biomarkers of tobacco smoke exposure in smokers.

Oxidative stress can contribute to the development of diseases, and may originate from exposures to toxicants commonly found in air pollution and cigarette smoke such as polycyclic aromatic hydrocarbons (PAHs) and volatile organic compounds (VOCs). Yet, associations between these exposures and oxidative stress biomarkers are poorly characterized. We report here novel associations between 14 exposure biomarkers of PAHs and VOCs, and two oxidative stress biomarkers; 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) and 8-isoprostaglandin F2α (8-isoprostane) in urine obtained from smokers participating in an ongoing clinical study (ESTxENDS, NCT03589989). We also assessed associations between six biomarkers of tobacco smoke exposure (metabolites of nicotine and tobacco-specific nitrosamines (TSNAs)) and both oxidative stress biomarkers. We then quantified the relative importance of each family of the 20 exposure biomarkers on oxidative stress. Participating smokers (153 men and 117 women, median age 44 years) had on average smoked 25 [2-62] years and smoked about 17 [5-40] cigarettes per day at the time of the study. Multiple linear regression results showed an association between 8-oxodG concentrations and the following metabolites in decreasing relative importance: PAHs (beta coefficient ß = 0.105, p-value <0.001, partial R2 = 0.15) > VOCs (ß = 0.028, p < 0.001, partial R2 = 0.09) > nicotine (ß = 0.226, p < 0.001, partial R2 = 0.08); and between 8-isoprostane concentrations and metabolites of PAHs (ß = 0.117, p < 0.001, partial R2 = 0.14) > VOCs (ß = 0.040, p < 0.001, partial R2 = 0.14) > TSNAs (ß = 0.202, p = 0.003, partial R2 = 0.09) > nicotine (ß = 0.266, p < 0.001, partial R2 = 0.08). Behavioral factors known to contribute to oxidative stress, including sleep quality, physical activity, and alcohol consumption, did not play a significant role. Exposures to PAHs and VOCs among smokers were significantly associated with oxidative stress.

Hand nicotine as an independent marker of thirdhand smoke pollution in children's environments.

BACKGROUND: Hand nicotine (HN) levels measure children's exposure to tobacco smoke pollutants from thirdhand and secondhand smoke. HN is associated with urinary and salivary cotinine, but the associations of HN with other tobacco smoke exposure (TSE) markers remain unknown. OBJECTIVES: We compared levels of HN and four urinary TSE biomarkers: cotinine, trans-3'-hydroxycotinine (3HC), nicotelline N-oxides, and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), and children's sociodemographic and TSE patterns. We also examined if HN is a plausible pathway for children's exposure to active smoking. METHODS: Data were collected from 175 non-smoking patients (Mean (SD) age = 5.4 (3.4) years) who lived with ≥1 cigarette smoker(s). HN and TSE biomarker levels were determined using LC-MS/MS. Multivariate and multivariable regression analyses were conducted to examine associations between TSE markers and parent-reported measures, controlling for sociodemographics. RESULTS: Of the five markers of TSE, cotinine (R2 = 0.221; p = 0.003) and HN (R2 = 0.247; p = 0.001) showed the strongest overall associations. Of the five markers, only cotinine showed significantly higher levels among Black children (ß^=0.307,p<0.05) independent of age, reported exposure, and home smoking bans. Cotinine (ß^=0.010,p<0.05), NNAL (ß^=0.012,p<0.05), and HN (ß^=0.011,p<0.05) showed significant positive associations with reported exposure independent of race, age, and home smoking bans. NNAL (ß^=-0.285,p<0.05) and HN (ß^=-0.336,p<0.05), but not cotinine, 3HC, and N-oxides, showed significantly lower levels among children who lived in homes with smoking bans. Child age, hand surface area, home smoking ban, and reported exposure independently accounted for 21 % of the variance in HN levels (p = 0.002). HN accounted for 30 % of the variance in cotinine independent of child race and child age. DISCUSSION: HN levels were associated with modifiable tobacco-related behaviors and shows promise as a marker of sources of THS pollution in a child's environment not captured by measurement of urinary cotinine alone. HN levels provide additional information about TSE, complementing other biomarkers when assessing children's overall TSE.

Optimum Urine Cotinine and NNAL Levels to Distinguish Smokers from Non-Smokers by the Changes in Tobacco Control Policy in Korea from 2008 to 2018.

INTRODUCTION: We examined the age- and sex-specific distributions of biomarkers of tobacco smoke exposure to determine the optimal cutoffs to distinguish smokers from non-smokers over the last 10 years in Korea, during which smoking prevalence and secondhand smoke (SHS) exposure declined due to changes in tobacco control policy. METHODS: We analyzed data from the Korea National Health and Nutrition Examination Survey on creatinine-adjusted urinary cotinine (2008-2018; 33 429 adults: 15 653 males and 17 776 females) and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL; 2016-2018; 6337 adults: 3091 males and 3246 females). We determined the optimal cutoffs and confidence intervals (CIs) to distinguish smokers from non-smokers using receiver operator characteristic curve analysis and bootstrapping (1000 resamples). RESULTS: The optimal cutoff values of creatinine-adjusted urine cotinine and NNAL concentration were 20.9 ng/mg (95% CI: 20.8-21.0, sensitivity: 96.6%, specificity: 93.8%) and 8.9 pg/mg (95% CI: 8.8-8.9, sensitivity: 94.0%, specificity: 94.7%), respectively, in 2016-2018. The optimal cutoffs of both biomarkers increased with age and were higher in females than in males for NNAL concentration. In both sexes, the optimal cutoff of urine cotinine continuously declined over the study period. CONCLUSIONS: The optimal cotinine cutoff declined along with smoking prevalence and levels of SHS exposure due to enforcement of tobacco control policies, including smoke-free ordinances and tax increases. Monitoring of biomarkers of tobacco exposure appears necessary for verification of smoking status and regulatory use. IMPLICATIONS: Our results based on nationally representative data suggest that a large decrease in the optimal cutoff value of urine cotinine to distinguish smokers from non-smokers was caused by decreases in smoking prevalence and SHS exposure following enforcement of tobacco control policies over the last 10 years. We determined the optimal cutoff values of urine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), which were not previously reported in representative population in Asia, to enable more accurate estimation of exposure to tobacco smoke and proper assessment of disease risks. Gender- and age-specific differences in the optimal cutoffs require further study. Monitoring of biomarkers of tobacco smoke exposure seems necessary for verification of smoking status and regulatory use.

Assessment of the Exposure to Tobacco-Specific Nitrosamines and Minor Tobacco Alkaloids in Users of Various Tobacco/Nicotine Products.

Tobacco-specific nitrosamines (TSNAs), in particular, the human carcinogens 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and N'-nitrosonornicotine (NNN), are important toxicants in tobacco and also (as contaminants) in nicotine products. In a clinical study comprising a period of 74 h under confinement, we investigated the exposure to NNK, NNN, N'-nitrosoanabasine (NAB), and N'-nitrosoanatabine (NAT) as well as to the minor tobacco alkaloids anabasine (AB) and anatabine (AT) by measuring suitable biomarkers in habitual users of combustible cigarettes (CCs), electronic cigarettes (ECs), heated tobacco products (HTPs), oral tobacco (OT), and nicotine replacement therapy products (NRTs). Non-users (NU) of any tobacco/nicotine products served as the (negative) control group. Smokers exhibited the highest levels for all biomarkers measured, except for AB in urine, which was found to be highest in OT users. Somewhat elevated levels compared to NU, EC, and NRT groups were also observed in the users of HTPs. In the users of tobacco-containing products (CC, HTP, and OT), most frequently the biomarkers significantly correlated with the dose markers such as daily consumption, urinary nicotine equivalents (Nequ), and plasma cotinine (CotP). In conclusion, except for smokers (CC) and OT users, exposure of users of ECs, HTPs, and NRTs to TSNAs as well as the minor tobacco alkaloids AB and AT is marginal and statistically not distinguishable from that of NU. Finally, our results for NNN in the saliva provide preliminary evidence for the formation of NNN from the precursor nornicotine in the presence of thiocyanate as a catalyst. The latter hypothesis requires experimental verification.

Assessment of the Exposure to NNN in the Plasma of Smokeless Tobacco Users.

N-Nitrosonornicotine (NNN) is a human carcinogen present in cigarette smoke and smokeless tobacco. Urinary NNN is usually measured in order to assess the exposure to this toxicant for tobacco users. NNN excretion in urine can be highly biased due to the formation of NNN by nitrosation of nornicotine under acidic conditions, both endogenously and exogenously. Hence, urinary NNN levels may not necessarily correctly reflect the product-specific exposure. Measurement of plasma NNN may be less prone to endogenous formation due to the stable pH (7.4) of blood. We developed an LC-MS/MS method for the quantification of NNN using 1 mL of human plasma. Validation according to FDA guidelines proved that the method is selective and highly sensitive with an LLOQ of 0.3 pg/mL. Accuracy and precision averaged to 98.7 and 7.5% (CV), respectively. The assay was applied to plasma samples collected from 10 experienced moist smokeless tobacco users during and after a single use of 2 g of the product for 40 min under controlled use conditions. Blood was drawn at 15 time points over a 6 h time course. The maximum NNN concentration (Cmax) ranged from 3.5 to 10 pg/mL (mean: 7.1 pg/mL) at a tmax of 32 min. Plasma NNN and nicotine were found to have similar time courses. In conclusion, the determination of NNN in plasma may be fit-for-purpose to evaluate the product-use-specific exposure to this carcinogen.

Simultaneous analysis of urinary total 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol, N'-nitrosonornicotine, and cotinine by liquid chromatography-tandem mass-spectrometry.

Biomarkers of exposure to harmful tobacco constituents are key tools for identifying individuals at risk and developing interventions and tobacco control measures. However, tobacco biomarker studies are scarce in many parts of the world with high prevalence of tobacco use. Our goal was to establish a robust method for simultaneous analysis of urinary total 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), N'-nitrosonornicotine (NNN), and cotinine at the Advanced Centre for Treatment, Research and Education in Cancer (ACTREC) in Mumbai, India. These biomarkers are validated measures of exposure to the carcinogenic tobacco nitrosamines 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and NNN and the addictive alkaloid nicotine, respectively. The established method is characterized by excellent accuracy, linearity, and precision, and was successfully applied to the analysis of 15 smokeless tobacco (SLT) users and 15 non-users of tobacco recruited in Mumbai. This is the first report of establishment of such procedure in a laboratory in India, which offers the first in-country capacity for research on tobacco carcinogenesis in Indian SLT users.

Effect of an electronic nicotine delivery system with 0, 8, or 36 mg/mL liquid nicotine versus a cigarette substitute on tobacco-related toxicant exposure: a four-arm, parallel-group, randomised, controlled trial.

BACKGROUND: Electronic nicotine delivery systems (ENDSs) are used by some smokers to reduce cigarette consumption, but their effectiveness is uncertain. We aimed to examine the extent to which ENDSs or a non-nicotine cigarette substitute influence tobacco-related toxicant exposure and cigarette consumption in smokers interested in smoking reduction. METHODS: We did a four-arm, parallel-group, randomised controlled trial at two sites in the USA (Penn State University, Hershey, PA, and Virginia Commonwealth University, Richmond, VA). We enrolled adults aged 21-65 years who smoked more than nine cigarettes per day (for at least the past year), with exhaled CO of more than 9 parts per million at screening, who were not currently using an ENDS, and who were interested in reducing smoking but not quitting. Participants were randomised (site-specific with allocation concealment; 1:1:1:1) to receive either a cartomiser-based, pen-style ENDS (eGo-style) paired with 0, 8, or 36 mg/mL liquid nicotine (participants and researchers masked to concentration) or a non-ENDS cigarette-shaped plastic tube that delivered no nicotine or aerosol (cigarette substitute; unmasked) for 24 weeks. Conditions were chosen to reflect a range of nicotine delivery including none (cigarette substitute and 0 mg/mL ENDS), low (8 mg/mL), and cigarette-like (36 mg/mL), and all conditions were paired with smoking reduction instructions. The primary outcome was concentration of the tobacco-specific carcinogen metabolite 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL; urinary total) collected at randomisation and at 4, 12, and 24 weeks. Multiple imputation with and without covariate adjustment was used in addition to sensitivity analyses. This trial is registered with ClinicalTrials.gov, NCT02342795. FINDINGS: Between July 22, 2015, and Nov 16, 2017, 684 individuals were screened and 520 (76%) were enrolled and randomised. 188 (36%) of 520 participants were lost to follow-up by week 24; attrition did not differ by study group (39 [30%] of 130 in the cigarette substitute group, 56 [43%] of 130 in the ENDS with 0 mg/mL nicotine group, 49 [38%] of 130 in the ENDS 8 mg/mL group, and 44 [34%] of 130 in the ENDS 36 mg/mL group). Urinary total NNAL at 24 weeks in the ENDS with 36 mg/mL nicotine group was 210·80 pg/mg creatinine (95% CI 163·03-274·42) compared with 346·09 pg/mg creatinine (265·00-455·32) in the cigarette substitute group (p=0·0061). No other significant differences between groups were observed for any time point for urinary total NNAL. Serious adverse event frequency was similar across groups (12 events in 12 participants [9%] in the ENDS with 36 mg/mL nicotine group, seven events in six participants [5%] in the 8 mg/mL group, 11 events in ten participants [8%] in the 0 mg/mL group, and 13 events in 13 participants [10%] in the cigarette substitute group), and all of these were deemed unrelated or unlikely to be related to study product use. There was one death between randomisation and 24 weeks (suicide; in the ENDS with 0 mg/mL nicotine group). INTERPRETATION: Use of an ENDS with cigarette-like nicotine delivery can reduce exposure to a major pulmonary carcinogen, NNAL, even with concurrent smoking. Future ENDS trials should involve products with well characterised nicotine delivery, including those with nicotine delivery approaching that of a cigarette. FUNDING: National Institutes of Health, US Food and Drug Administration.

Association between Second-hand Smoke Exposure and Urinary NNAL Level in Korean Adolescents.

BACKGROUND: The 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) is a metabolite of tobacco-specific lung carcinogen that can be found in both smokers and non-smokers. Particularly, NNAL levels of children with a history of exposure to second-hand smoke (SHS) are higher than those of adults. Thus, we aimed to investigate the association between SHS exposure and urine NNAL levels in Korean adolescents. METHODS: This cross-sectional study used data from the Korea National Health and Nutrition Examination Survey VII. Overall, 648 never-smoking adolescents (425 boys and 223 girls) aged 12 to 18 were included in this study. Logistic regression analyses identified the relationship between SHS exposure and elevated urine NNAL levels. RESULTS: The mean urine NNAL levels of the no exposure and exposure group in boys were 1.39 and 2.26 ng/mL, respectively, whereas they were 1.01 and 2.45 ng/mL in girls, respectively (P < 0.001). Among the adolescents exposed to SHS, the confounder-adjusted odds ratio (95% confidence intervals) for elevated urine NNAL levels according to exposure area as overall, home, and public area were 2.68 (1.58-4.53), 31.02 (9.46-101.74), and 1.89 (1.12-3.17) in boys; and 6.50 (3.22-13.11), 20.09 (7.08-57.04), and 3.94 (1.98-7.77) in girls, respectively. CONCLUSION: SHS exposure was significantly associated with elevated urine NNAL levels in Korean adolescents, particularly in female adolescents and in those with home exposure. These findings remind us of the need to protect adolescents from SHS.

Association of a Tobacco-specific Nitrosamine Carcinogen with Urinary Cotinine, Urinary Sodium Excretion, and Total Energy Intake in Adolescents and Children.

This study investigated the association of a tobacco-specific nitrosamine carcinogen, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) with urinary cotinine (uCot), urinary sodium (uNa) excretion, systolic blood pressure (sBP), and total energy intake in adolescents and children in relation to the subjects' age. A total of 790 subjects aged 6-19 years were evaluated. NNAL, uCot, corrected NNAL (cNNAL), the NNAL/uCot ratio, uNa, sBP, and nutrient intake were measured. A strong association between uCot and cNNAL was observed in children who were 11 years of age (r=0.881, P<0.001); however, no significant association was noted in adolescents who were 19 years of age. The uNa level was significantly higher (133.9 mmol/L vs. 107.8 mmol/L, P<0.001) and sBP was significantly lower (105.3 mmHg vs. 110.6 mmHg, P=0.012) in adolescents with elevated NNAL than in those without elevated NNAL. NNAL was significantly higher in subjects with increased uNa excretion than in those without increased uNa excretion. NNAL was positively correlated with uNa (r=0.183, P<0.001) and negatively correlated with sBP (r=-0.142, P<0.001). Non-smokers with elevated NNAL/uCot ratios had significantly lower total energy intake than those without elevated NNAL/uCot ratios (1729.0 kcal/day vs. 1911.0 kcal/day, P=0.008). The relationship between NNAL and uCot varied according to the subjects' age. NNAL seems to play a role in decreasing sBP by enhancing uNa excretion. Insufficient nutrient intake may contribute to endogenous formation of NNAL in non-smoking adolescents and children.

Concentrations of Cotinine and 4-(Methylnitrosamino)-1-(3-Pyridyl)-1-Butanol (NNAL) in U.S. Non-Daily Cigarette Smokers.

BACKGROUND: Accumulating evidence suggests that non-daily smokers have higher disease and mortality risks than never smokers. Yet, the accuracy of self-reported non-daily cigarette smoking is poorly understood. METHODS: We examined the concordance between self-reported non-daily smoking and serum cotinine in 18,835 adult participants (20 years or older) of the 2007 to 2014 National Health and Nutrition Examination Surveys, in comparison with daily smokers and nonsmokers. We also analyzed concentrations of the urinary biomarker 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) by smoking status. RESULTS: In the study sample, 77.8% (14,660) reported currently not smoking (nonsmokers), 18.3% (3,446) smoked every day (daily smokers), and 3.9% (729) smoked on some days of the past month (non-daily smokers). Just 2.1% of nonsmokers had cotinine concentrations in the active smoking range (>10 ng/mL), compared with 70.4% of non-daily and 98.8% of daily smokers. Non-daily smokers reported smoking a median of 24 cigarettes per month [interquartile range (IQR) = 9-60] and had substantially higher concentrations of NNAL (median = 72.5; IQR = 14.8-211.0 pg/mL) than nonsmokers (median = 0.4; IQR = 0.4-2.1 pg/mL), although lower than daily smokers (median = 294.0; IQR = 148.0-542.0 pg/mL). Among non-daily smokers, concentrations of cotinine and NNAL were positively correlated with days and cigarettes smoked per month (P < 0.001). CONCLUSIONS: We observed excellent concordance between self-reported non-daily cigarette smoking and concentrations of serum cotinine. IMPACT: These results provide evidence for the validity of self-reported non-daily smoking and indicate that non-daily smokers are exposed to substantial concentrations of carcinogenic nitrosamines regardless of the low number of cigarettes they smoke per month.

Exposure to Nicotine and Toxicants Among Dual Users of Tobacco Cigarettes and E-Cigarettes: Population Assessment of Tobacco and Health (PATH) Study, 2013-2014.

INTRODUCTION: Concurrent use of tobacco cigarettes and e-cigarettes ("dual use") is common among tobacco users. Little is known about differences in demographics and toxicant exposure among subsets of dual users. AIMS AND METHODS: We analyzed data from adult dual users (current every/some day users of tobacco cigarettes and e-cigarettes, n = 792) included in the PATH Study Wave 1 (2013-2014) and provided urine samples. Samples were analyzed for biomarkers of exposure to nicotine and selected toxicants (tobacco-specific nitrosamine NNK [NNAL], lead, cadmium, naphthalene [2-naphthol], pyrene [1-hydroxypyrene], acrylonitrile [CYMA], acrolein [CEMA], and acrylamide [AAMA]). Subsets of dual users were compared on demographic, behavioral, and biomarker measures to exclusive cigarette smokers (n = 2411) and exclusive e-cigarette users (n = 247). RESULTS: Most dual users were predominant cigarette smokers (70%), followed by daily dual users (13%), non-daily concurrent dual users (10%), and predominant vapers (7%). Dual users who smoked daily showed significantly higher biomarker concentrations compared with those who did not smoke daily. Patterns of e-cigarette use had little effect on toxicant exposure. Dual users with high toxicant exposure were generally older, female, and smoked more cigarettes per day. Dual users who had low levels of biomarkers of exposure were generally younger, male, and smoked non-daily. CONCLUSIONS: In 2013-2014, most dual users smoked cigarettes daily and used e-cigarettes occasionally. Cigarette smoking appears to be the primary driver of toxicant exposure among dual users, with little-to-no effect of e-cigarette use on biomarker levels. Results reinforce the need for dual users to stop smoking tobacco cigarettes to reduce toxicant exposure. IMPLICATIONS: With considerable dual use of tobacco cigarettes and e-cigarettes in the United States, it is important to understand differences in toxicant exposure among subsets of dual users, and how these differences align with user demographics. Findings suggest most dual users smoke daily and use e-cigarettes intermittently. Low exposure to toxicants was most common among younger users, males, and intermittent smokers; high exposure to toxicants was most common among older users, females, and heavier cigarette smokers. Results underscore the heterogeneity occurring within dual users, and the need to quit smoking cigarettes completely in order to reduce toxicant exposure.

Does the implementation of smoke-free laws and smoking culture affect exposure to tobacco smoking? Results from 3 hospitality settings in South Korea.

OBJECTIVES: It is especially difficult for hospitality workers to avoid secondhand smoke (SHS), meaning that they are likely particularly vulnerable to the effects of SHS. The authors aimed to determine the degree to which smoke-free laws protect hospitality workers from SHS exposure, by examining biochemical markers of such exposure. MATERIAL AND METHODS: This was a cross-sectional study examining SHS exposure in non-smoking employees working in hospitality settings where smoking is prohibited or permitted. The following biomarkers were selected: cotinine and tobaccospecific nitrosamines, which are known to measure SHS exposure, and 2 representative carcinogens: 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK). The authors compared these biomarkers between 3 hospitality settings. A descriptive analysis was performed. In addition, they conducted 1-way and 2-way analysis of covariance (ANCOVA) to compare the biochemical markers. RESULTS: Smoking substances were identified by smoking ban levels. In the case of hair nicotine and urine cotinine, their concentrations were lower in areas with a complete smoking ban than in both areas with a separate smoking room and no smoking ban; however, there was no statistically significant difference between these. In the case of dust NNK, its level was the lowest in areas with a complete smoking ban. To confirm the smoking ban effect by hospitality settings, the authors checked the results of the 2-way ANCOVA. In karaoke and billiard halls, the dust NNK concentrations were significantly higher in areas with no smoking ban than in areas with a separate smoking room. CONCLUSIONS: Exposure to SHS is more prevalent in places that are more lenient when it comes to smoking (e.g., Internet cafés) than in places that are not (e.g., restaurants and cafés), even when smoking is similarly prohibited in both types of places. Int J Occup Med Environ Health. 2021;34(1):53-67.

Tobacco-Specific Nitrosamines (NNAL, NNN, NAT, and NAB) Exposures in the US Population Assessment of Tobacco and Health (PATH) Study Wave 1 (2013-2014).

INTRODUCTION: The tobacco-specific nitrosamines (TSNAs) are an important group of carcinogens found in tobacco and tobacco smoke. To describe and characterize the levels of TSNAs in the Population Assessment of Tobacco and Health (PATH) Study Wave 1 (2013-2014), we present four biomarkers of TSNA exposure: N'-nitrosonornicotine, N'-nitrosoanabasine, N'-nitrosoanatabine, and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) which is the primary urinary metabolite of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone. METHODS: We measured total TSNAs in 11 522 adults who provided urine using automated solid-phase extraction coupled to isotope dilution liquid chromatography-tandem mass spectrometry. After exclusions in this current analysis, we selected 11 004 NNAL results, 10 753 N'-nitrosonornicotine results, 10 919 N'-nitrosoanatabine results, and 10 996 N'-nitrosoanabasine results for data analysis. Geometric means and correlations were calculated using SAS and SUDAAN. RESULTS: TSNA concentrations were associated with choice of tobacco product and frequency of use. Among established, every day, exclusive tobacco product users, the geometric mean urinary NNAL concentration was highest for smokeless tobacco users (993.3; 95% confidence interval [CI: 839.2, 1147.3] ng/g creatinine), followed by all types of combustible tobacco product users (285.4; 95% CI: [267.9, 303.0] ng/g creatinine), poly tobacco users (278.6; 95% CI: [254.9, 302.2] ng/g creatinine), and e-cigarette product users (6.3; 95% CI: [4.7, 7.9] ng/g creatinine). TSNA concentrations were higher in every day users than in intermittent users for all the tobacco product groups. Among single product users, exposure to TSNAs differed by sex, age, race/ethnicity, and education. Urinary TSNAs and nicotine metabolite biomarkers were also highly correlated. CONCLUSIONS: We have provided PATH Study estimates of TSNA exposure among US adult users of a variety of tobacco products. These data can inform future tobacco product and human exposure evaluations and related regulatory activities.