The objective of this study was to examine long-term trends in serum cotinine (COT) concentrations, as a measure of secondhand smoke (SHS) exposure, in U.S. nonsmokers using data from the National Health and Nutrition Examination Surveys (NHANES) from 2003 to 2018. We analyzed NHANES serum COT results from 8 continuous NHANES 2 year cycles from 2003 to 2018 using a liquid chromatography–tandem mass spectrometry assay that has been maintained continuously at the Centers for Disease Control and Prevention (CDC) since 1992. Serum COT concentrations (based on the geometric means) among nonsmokers in the U.S. decreased by an average of 11.0% (95% confidence interval (CI) [8.8%, 13.1%]; p < 0.0001) every 2 year cycle. From 2003 to 2018, serum COT concentrations in U.S. nonsmokers declined by 55.0%, from 0.065 ng/mL in 2003–2004 to 0.029 ng/mL in 2017–2018 (p < 0.0001). Significant decreases in serum COT concentrations were observed in all demographic groups. While disparities between these groups seems to be shrinking over time, several previously observed disparities in SHS exposure remain in 2017–2018. Serum COT concentrations of the non-Hispanic Black population remained higher than those of non-Hispanic Whites and Mexican Americans (p < 0.0001). Additionally, serum COT concentrations were significantly higher for children aged 3–5 years than other age groups (p ≤ 0.0002), and men continued to have significantly higher serum COT concentrations than women (p = 0.0384). While there is no safe level of exposure to SHS, the decrease in serum COT concentrations in the U.S. population as well as across demographic groupings represents a positive public health outcome and supports the importance of comprehensive smoke-free laws and policies for workplaces, public places, homes, and vehicles to protect nonsmokers from SHS exposure. © 2022 by the authors. Licensee MDPI, Basel, Switzerland.

INTRODUCTION: The Population Assessment of Tobacco and Health (PATH) Study is a longitudinal cohort study on tobacco use behavior, attitudes and beliefs, and tobacco-related health outcomes, including biomarkers of tobacco exposure in the U.S. population. In this report we provide a summary of urinary nicotine metabolite measurements among adult users and non-users of tobacco from Wave 1 (2013-2014) of the PATH Study. METHODS: Total nicotine and its metabolites including cotinine, trans-3'-hydroxycotinine (HCTT), and other minor metabolites were measured in more than 11 500 adult participants by liquid chromatography tandem mass spectrometry methods. Weighted geometric means (GM) and least square means from statistical modeling were calculated for non-users and users of various tobacco products. RESULTS: Among daily users, the highest GM concentrations of nicotine, cotinine and HCTT were found in exclusive smokeless tobacco users, and the lowest in exclusive e-cigarette users. Exclusive combustible product users had intermediate concentrations, similar to those found in users of multiple products (polyusers). Concentrations increased with age within the categories of tobacco users, and differences associated with gender, race/ethnicity and educational attainment were also noted among user categories. Recent (past 12 months) former users had GM cotinine concentrations that were more than threefold greater than never users. CONCLUSIONS: These urinary nicotine metabolite data provide quantification of nicotine exposure representative of the entire US adult population during 2013-2014 and may serve as a reference for similar analyses in future measurements within this study. IMPLICATIONS: Nicotine and its metabolites in urine provide perhaps the most fundamental biomarkers of recent nicotine exposure. This report, based on Wave 1 of the Population Assessment of Tobacco and Health (PATH) Study, provides the first nationally representative data describing urinary nicotine biomarker concentrations in both non-users, and users of a variety of tobacco products including combustible, e-cigarette and smokeless products. These data provide a urinary biomarker concentration snapshot in time for the entire US population during 2013-2014, and will provide a basis for comparison with future results from continuing, periodic evaluations in the PATH Study.

INTRODUCTION: The Population Assessment of Tobacco and Health (PATH) Study is a nationally representative cohort of tobacco product users and nonusers. The study's main purpose is to obtain longitudinal epidemiologic data on tobacco use and exposure among the US population. AIMS AND METHODS: Nicotine biomarkers-cotinine (COT) and trans-3'-hydroxycotinine (HCT)-were measured in blood samples collected from adult daily tobacco users and nonusers during Wave 1 of the PATH Study (2013-2014; n = 5012; one sample per participant). Participants' tobacco product use and exposure to secondhand smoke were categorized based on questionnaire responses. Nonusers were subdivided into never users and recent former users. Daily tobacco users were classified into seven tobacco product use categories: exclusive users of cigarette, smokeless tobacco, electronic cigarette, cigar, pipe, and hookah, as well as polyusers. We calculated sample-weighted geometric mean (GM) concentrations of cotinine, HCT, and the nicotine metabolite ratio (NMR) and evaluated their associations with tobacco use with adjustment for potential confounders. RESULTS: The GMs (95% confidence intervals) of COT and HCT concentrations for daily tobacco users were 196 (184 to 208) and 72.5 (67.8 to 77.4) ng/mL, and for nonusers they were 0.033 (0.028 to 0.037) and 0.021 (0.018 to 0.023) ng/mL. Exclusive smokeless tobacco users had the highest COT concentrations of all user groups examined. The GM NMR in daily users was 0.339 (95% confidence interval: 0.330 to 0.350). CONCLUSIONS: These nationally representative estimates of serum nicotine biomarkers could be the basis for reference ranges characterizing nicotine exposure for daily tobacco users and nonusers in the US adult population. IMPLICATIONS: This report summarizes the serum nicotine biomarker measurements in Wave 1 of the PATH Study. We are reporting the first estimates of HCT in serum for daily tobacco users and nonusers in the noninstitutionalized, civilian US adult population; the first nationally representative serum COT estimates for daily exclusive users of different tobacco products and daily polyusers; and the first nationally representative estimate of the serum NMR in daily tobacco users by age, race/ethnicity, and sex.

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.

BACKGROUND: The changing prevalence and patterns of tobacco use, the advent of novel nicotine delivery devices, and the development of new biomarkers prompted an update of the 2002 Society for Research on Nicotine and Tobacco (SRNT) report on whether and how to apply biomarker verification for tobacco use and abstinence. METHODS: The SRNT Treatment Research Network convened a group of investigators with expertise in tobacco biomarkers to update the recommendations of the 2002 SNRT Biochemical Verification Report. RESULTS: Biochemical verification of tobacco use and abstinence increases scientific rigor and is recommended in clinical trials of smoking cessation, when feasible. Sources, appropriate biospecimens, cutpoints, time of detection windows and analytic methods for carbon monoxide, cotinine (including over the counter tests), total nicotine equivalents, minor tobacco alkaloids, and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol are reviewed, as well as biochemical approaches to distinguishing cigarette smoking from use of electronic nicotine delivery devices (ENDS). CONCLUSIONS: Recommendations are provided for whether and how to use biochemical verification of tobacco use and abstinence. Guidelines are provided on which biomarkers to use, which biospecimens to use, optimal cutpoints, time windows to detection, and methodology for biochemical verifications. Use of combinations of biomarkers is recommended for assessment of ENDS use. IMPLICATIONS: Biochemical verification increases scientific rigor, but there are drawbacks that need to be assessed to determine whether the benefits of biochemical verification outweigh the costs, including the cost of the assays, the feasibility of sample collection, the ability to draw clear conclusions based on the duration of abstinence, and the variability of the assay within the study population. This paper provides updated recommendations from the 2002 SRNT report on whether and how to use biochemical markers in determining tobacco use and abstinence.

Urinary cotinine is one of the most commonly measured biomarkers reflecting recent exposure to nicotine. In some cases a simple qualitative dichotomization of smokers and non-smokers is all that is required. NicAlert(R) test strips have been evaluated for this purpose, but other recently introduced, inexpensive single-line test strips have not. In this study we evaluated two such strips with nominal cutoffs of 200 and 10 ng/mL. A total of 800 urine samples with known cotinine concentrations determined by an LC-MS-MS method were examined, including 400 urine samples ranging from 0.23 to more than 24,000 ng/mL by the 200 ng/mL strip, and 400 samples with concentrations <200 ng/mL by the 10 ng/mL cutoff strip. Both test strips performed well in these evaluations. Classification relative to LC-MS-MS by the 200 ng/mL strips had a sensitivity of 99.5% and specificity of 92%, with 95.8% accuracy. The 10 ng/mL strips had a sensitivity of 98.7% and specificity of 90.1%, with 93.3% accuracy. The positive predictive value for the 200 ng/mL strips was 92.6% and the negative predictive value was 99.5%. For the 10 ng/mL strips, the corresponding values were 85.4 and 99.2%, respectively. The prevalence of positive samples was 50% in the 200 ng/mL group, and 37% in the 10 ng/mL set. Each strip was read by two readers with an overall agreement of >98%. Our results suggest that these simple and inexpensive lateral flow immunoassay test strips can provide useful qualitative estimates of nicotine exposures for appropriate applications within the inherent limitations of sensitivity and precision of the immunoassay test strip format.

BACKGROUND: Biomarkers of tobacco exposure have a central role in studies of tobacco use and nicotine intake. The most significant exposure markers are nicotine itself and its metabolites in urine. Therefore, it is important to evaluate the performance of laboratories conducting these biomarker measurements. METHODS: This report presents the results from a method performance study involving 11 laboratories from 6 countries which are currently active in this area. Each laboratory assayed blind replicates of 7 human urine pools at various concentrations on 3 separate days. The samples included 5 pools blended from smoker and nonsmoker urine sources, and 2 additional blank urine samples fortified with pure nicotine, cotinine and hydroxycotinine standards. All laboratories used their own methods, and all were based on some form of liquid chromatography / tandem mass spectrometry. RESULTS: Overall, good agreement was found among the laboratories in this study. Intralaboratory precision was good, and in the fortified pools the mean bias observed was &lt; + 3.5% for nicotine, approximately 1.2% for hydroxycotinine, and less than 1% for cotinine (1 outlier excluded in each case). Both indirect and direct methods for analyzing the glucuronides gave comparable results. CONCLUSIONS: This evaluation indicates that the experienced laboratories participating in this study can produce reliable and comparable human urinary nicotine metabolic profiles in samples from people with significant recent exposure to nicotine. IMPACT: This work supports the reliability and agreement of an international group of established laboratories measuring nicotine and its metabolites in urine in support of nicotine exposure studies.

Accurate and reliable measurements of exposure to tobacco products are essential for identifying and confirming patterns of tobacco product use and for assessing their potential biological effects in both human populations and experimental systems. Due to the introduction of new tobacco-derived products and the development of novel ways to modify and use conventional tobacco products, precise and specific assessments of exposure to tobacco are now more important than ever. Biomarkers that were developed and validated to measure exposure to cigarettes are being evaluated to assess their utility for measuring exposure to these new products. Here, we review current methods for measuring exposure to new and emerging tobacco products, such as electronic cigarettes, little cigars, water pipe and cigarillos. Rigorously validated biomarkers specific to these new products are yet to be identified. Here, we discuss the strengths and limitations of current approaches, including whether or not they provide reliable exposure estimates. We provide specific guidance for choosing practical and economical biomarkers for different study designs and experimental conditions. Our goal is to help both new and experienced investigators measure exposure to tobacco products accurately, while avoiding common experimental errors. By identifying the capacity gaps in biomarker research on new and emerging tobacco products, we hope to provide researchers, policy makers and funding agencies with a clear action plan for conducting and promoting research on the patterns of use and health effects of these products.

BACKGROUND: The workplace is one of the major locations outside of the home for nonsmokers' exposure to secondhand smoke (SHS). New policies in many states and localities restrict or prohibit smoking in the worksites and information on current trends in the exposure of nonsmokers to SHS across various occupational groups is therefore needed. OBJECTIVE: To evaluate temporal trends in SHS exposure among nonsmoking workers in the United States, and identify those occupations with workers with the highest levels of SHS exposure. METHODS: We combined serum cotinine (sCOT) measurements and questionnaire data from five survey cycles of the National Health and Nutrition Examination Survey (NHANES: 2001-2010). Trends of SHS exposure by occupations were examined by percent changes and least-squares geometric means (LSGMS) of sCOT concentrations computed using sample-weighted multiple regression models. RESULTS: Between NHANES 2001-02 and 2009-10, LSGMs of sCOT levels had changed -25% (95% CI: -39, -7%) in nonsmoking workers. The largest decrease was identified among food preparation workers -54% (95% CI: -74, -19%), followed by white collar (-40%, 95% CI: -56, -19%) and blue collar workers (-32%, 95% CI: -51, -5%). LSGMs of sCOT remained highest in food preparation workers in all survey cycles, but the gap between occupations narrowed in the latest survey cycle (2009-10). For instance, the gap in LSGMs of sCOT between food preparation and science/education workers dropped above 70% during 2000 to 2010. CONCLUSIONS: During the period from 2001 to 2010, the overall SHS exposure in nonsmoking workers has declined with substantial decline in food preparation/service and blue-collar workers. Although disparities persist in SHS exposure, the gap among occupations has narrowed.

OBJECTIVE: To estimate changes in nicotine intakes in cigarette smokers in the United States from 1988 to 2012 using the National Health and Nutrition Examination Survey (NHANES). METHODS: NHANES provides data on nationally representative samples of cigarette smokers from the civilian noninstitutionalized U.S. population. A total of 4,304 smokers aged 20 and above were studied in NHANES III 1988-1994 and 7,095 in the continuous NHANES 1999-2012. We examined serum cotinine concentrations, daily cigarette consumption, and estimated nicotine intake per cigarette, with adjustment for sex, age, racial/ethnic background, education level, and body mass index. RESULTS: There was little overall change in nicotine intake from smoking cigarettes either in the U.S. population as a whole or in major racial/ethnic subgroups over the 25-year period from 1988. Serum cotinine averaged 223.7ng/mL (95% confidence interval [CI] = 216.1-231.3) in 1988-1994, which was not significantly different from the adjusted mean of 219.2ng/mL (95% CI = 214.1-224.4) in 1999-2012. Over the same period, average daily cigarette consumption declined substantially, from 17.3 (95% CI = 16.5-18.0) in 1988-1994 to 12.3 (95% CI = 11.0-13.6) by 2012. Cotinine per cigarette smoked increased by some 42% between 1988-1994 and 2011-2012, from a geometric mean of 12.4 (95% CI = 11.7-13.1) to 17.6 (95% CI = 16.1-19.2). CONCLUSIONS: Reductions in cigarette smoking prevalence since the late 1980s, changes in cigarette product design, and the widespread introduction of smoke-free policies have not impacted significantly on nicotine intakes in U.S. smokers. Reductions in cigarette consumption have been offset by increased nicotine intake per cigarette smoked.

Tobacco use is a major contributor to premature morbidity and mortality. The measurement of nicotine and its metabolites in urine is a valuable tool for evaluating nicotine exposure and for nicotine metabolic profiling-i.e., metabolite ratios. In addition, the minor tobacco alkaloids-anabasine and anatabine-can be useful for monitoring compliance in smoking cessation programs that use nicotine replacement therapy. Because of an increasing demand for the measurement of urinary nicotine metabolites, we developed a rapid, low-cost method that uses isotope dilution liquid chromatography-tandem mass spectrometry (LC-MS/MS) for simultaneously quantifying nicotine, six nicotine metabolites, and two minor tobacco alkaloids in smokers' urine. This method enzymatically hydrolyzes conjugated nicotine (primarily glucuronides) and its metabolites. We then use acetone pretreatment to precipitate matrix components (endogenous proteins, salts, phospholipids, and exogenous enzyme) that may interfere with LC-MS/MS analysis. Subsequently, analytes (nicotine, cotinine, hydroxycotinine, norcotinine, nornicotine, cotinine N-oxide, nicotine 1'-N-oxide, anatabine, and anabasine) are chromatographically resolved within a cycle time of 13.5 minutes. The optimized assay produces linear responses across the analyte concentrations typically found in urine collected from daily smokers. Because matrix ion suppression may influence accuracy, we include a discussion of conventions employed in this procedure to minimize matrix interferences. Simplicity, low cost, low maintenance combined with high mean metabolite recovery (76-99%), specificity, accuracy (0-10% bias) and reproducibility (2-9% C.V.) make this method ideal for large high through-put studies.

BACKGROUND: Tobacco-specific carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) was measured in all participants aged 6 years and older from the Centers for Disease Control and Prevention's National Health and Nutrition Examination Survey 2007-2008. The suitability of using creatinine or specific gravity for urinary NNAL correction in exposure assessment is examined in this study. METHODS: Effects of both specific gravity and creatinine correction on urinary NNAL among smokers were investigated with multiple linear regression models using either normalization or the fitting of creatinine and specific gravity in the model as covariates. RESULTS: When log-scaled NNAL was normalized by either creatinine or specific gravity, R2 was slightly higher for creatinine than for specific gravity (R2 = 0.1694 and 0.1439, for creatinine and specific gravity, respectively). When log-scaled NNAL was normalized by both factors, the R2 was improved (R2 = 0.2068). When specific gravity or creatinine was included as a covariate separately in the models, they were highly significant factors (P < 0.001, R2 = 0.2226 and 0.1681 for creatinine and specific gravity, respectively). However, when both were included in the model as covariates, creatinine remained highly significant (P < 0.001), whereas the significance of specific gravity was eliminated (P = 0.4294). CONCLUSION: This study confirms significant relationships between NNAL concentrations and both urine creatinine and specific gravity. We conclude that creatinine is the more influential and preferred variable to account for urine dilution in tobacco-specific nitrosamine exposure assessment.

We report on the results of a low-intensity behavioral intervention to reduce second hand smoke (SHS) exposure of children with asthma from low income minority households in Los Angeles, California. In this study, 242 child/adult dyads were randomized to a behavioral intervention (video, workbook, minimal counseling) or control condition (brochure). Main outcome measures included child's urine cotinine and parental reports of child's hours of SHS exposure and number of household cigarettes smoked. Implementation of household bans was also considered. No differences in outcomes were detected between intervention and control groups at follow-up. Limitations included high attrition and low rates of collection of objective measures (few children with urine cotinine samples). There continues to be a need for effective culturally and linguistically appropriate strategies that support reduction of household SHS exposure among children with asthma in low income, minority households.

BACKGROUND: With an increase in indoor smoking bans, many smokers smoke outside establishments and near their entrances, which has become a public health concern. OBJECTIVES: We characterized the exposure of nonsmokers to secondhand smoke (SHS) outside a restaurant and bar in Athens, Georgia, where indoor smoking is banned, using salivary cotinine and urinary 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL).Methods: In a crossover study, we assigned 28 participants to outdoor patios of a restaurant and a bar and an open-air site with no smokers on three weekend days; participants visited each site once and stayed for 3 hr. We collected saliva and urine samples immediately before and after the visits (postexposure) and on the following morning and analyzed samples for cotinine and total NNAL, respectively. Regression models were fitted and changes in biomarkers were contrasted between locations. RESULTS: Postexposure and preexposure geometric mean salivary cotinine concentrations differed by 0.115 ng/mL [95% confidence interval (CI): 0.105, 0.126)] and by 0.030 ng/mL (95% CI: 0.028, 0.031) for bar and restaurant visits, respectively. There were no significant post- and preexposure differences in cotinine levels after control site visits, and changes after bar and restaurant site visits were significantly different from changes after control site visits (p < 0.001). Results comparing next-day and preexposure salivary cotinine levels were similar. Next-day creatinine-corrected urinary NNAL concentrations also were higher than preexposure levels following bar and restaurant visits [1.858 pg/mg creatinine higher (95% CI: 0.897, 3.758) and 0.615 pg/mg creatinine higher (95% CI: 0.210, 1.761), respectively], and were significantly different from changes after the control visits (p = 0.005). CONCLUSION: Salivary cotinine and urinary NNAL increased significantly in nonsmokers after outdoor SHS exposure. Our findings indicate that such exposures may increase risks of health effects associated with tobacco carcinogens.

OBJECTIVE: To test an air cleaner and health coach intervention to reduce secondhand smoke exposure compared with air cleaners alone or no air cleaners in reducing particulate matter (PM), air nicotine, and urine cotinine concentrations and increasing symptom-free days in children with asthma residing with a smoker. DESIGN: Randomized controlled trial, with randomization embedded in study database. SETTINGS: The Johns Hopkins Hospital Children's Center and homes of children. PARTICIPANTS: Children with asthma, residing with a smoker, randomly assigned to interventions consisting of air cleaners only (n = 41), air cleaners plus a health coach (n = 41), or delayed air cleaner (control) (n = 44). MAIN OUTCOME MEASURES: Changes in PM, air nicotine, and urine cotinine concentrations and symptom-free days during the 6-month study. RESULTS: The overall follow-up rate was high (91.3%). Changes in mean fine and coarse PM (PM(2.5) and PM(2.5-10)) concentrations (baseline to 6 months) were significantly lower in both air cleaner groups compared with the control group (mean differences for PM(2.5) concentrations: control, 3.5 mcg/m(3); air cleaner only, -19.9 mcg/m(3); and air cleaner plus health coach, -16.1 mcg/m(3); P = .003; and PM(2.5-10) concentrations: control, 2.4 mcg/m(3); air cleaner only, -8.7 mcg/m(3); and air cleaner plus health coach, -10.6 mcg/m(3); P = .02). No differences were noted in air nicotine or urine cotinine concentrations. The health coach provided no additional reduction in PM concentrations. Symptom-free days were significantly reduced in both air cleaner groups compared with the control group (P = .03). CONCLUSION: Although the use of air cleaners can result in a significant reduction in indoor PM concentrations and a significant increase in symptom-free days, it is not enough to prevent exposure to secondhand smoke. Trial Registration clinicaltrials.gov Identifier: NCT00466024.

Nicotine (NIC), cotinine (COT) and trans-3'-hydroxycotinine (OHCOT) are the most prevalent and abundant tobacco biomarkers in meconium. We have developed and validated an accurate and precise method for the measurement of these analytes in meconium in which potassium hydroxide is used to digest the meconium sample, followed by solid phase extraction from the liquified sample. The precision of OHCOT, COT and NIC measurements (intra-day and inter-day) were 4.8-10.6%, 3.4-11.6% and 9.3-15.8%, respectively. Evaluation of accuracy indicated bias of -4.0, 2.0 and 0.8% for OHCOT at concentrations of 0.5, 2.5 and 7.5ng/g. The accuracy estimates for COT at concentrations of 0.5, 2.5 and 7.5ng/g are 4.0, 4.0 and 5.7%, respectively. For NIC at 2, 10 and 30ng/g the accuracy was calculated to be 3.0, 5.0 and 5.1%, respectively. The linear range of standard solutions was 0.125-37.5ng/mL for OHCOT and COT, and 0.75-150ng/mL for NIC. This method was applied to the analysis of 374 meconium samples from infants of both smoking and nonsmoking mothers. Positive correlations with r(2)≥0.63 were observed between NIC and COT, COT and OHCOT, NIC and OHCOT, and NIC and (OHCOT+COT) in these samples.

OBJECTIVE: This study quantified casino dealers' occupational exposure to environmental tobacco smoke (ETS). METHODS: We measured casino dealers' exposure to ETS components by analyzing full-shift air and preshift and postshift urine samples. RESULTS: Casino dealers were exposed to nicotine, 4-vinyl pyridine, benzene, toluene, naphthalene, formaldehyde, acetaldehyde, solanesol, and respirable suspended particulates. Levels of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) in urine increased significantly during an 8-hour work shift both with and without adjustment for creatinine clearance. Creatinine-unadjusted cotinine significantly increased during the 8-hour shift, but creatinine-adjusted cotinine did not increase significantly. CONCLUSIONS: Casino dealers at the three casinos were exposed to airborne ETS components and absorbed an ETS-specific component into their bodies, as demonstrated by detectable levels of urinary NNAL. The casinos should ban smoking on their premises and offer employee smoking cessation programs.

The aromatic amine 4-aminobiphenyl (4-ABP) is present in tobacco smoke. In humans, it is also a known bladder carcinogen. We describe here a method for the quantification of total 4-ABP in urine using capillary gas chromatography/tandem mass spectrometry, with an effective detection limit in urine samples of approximately 0.87 pg/mL. We also examined the efficiency of chemical or enzymatic hydrolysis of urinary aromatic amine metabolites. Although we found acidic or basic hydrolysis effective, we found enzymatic hydrolysis (beta-glucuronidase with either Escherichia coli or Helix pomatia) ineffective. As part of this work, we also confirm the presence of N-acetyl-4-ABP and 4-ABP glucuronide in human urine samples from smokers. These metabolites have been reported in animal studies, but previously they have not been identified in human samples. These metabolites, however, were found to be unstable and thus infeasible for biomonitoring. The final validated urinary total 4-ABP assay was applied to the analysis of samples from smokers and nonsmokers, whose status was confirmed from cotinine EIA measurements. Among 41 confirmed nonsmokers, the geometric mean (95% CI) of 4-ABP concentration was 1.64 pg/mg creatinine (1.30-2.07). Conversely, in 89 smokers, the geometric mean of 4-ABP concentration was significantly greater, at 8.69 pg/mg creatinine (7.43-10.16), p < 0.001. Our results indicate that following tobacco smoke exposure, total urinary 4-ABP is a reliable biomarker for exposure to this carcinogen.

BACKGROUND: The Food and Drug Administration (FDA) is examining options for regulating menthol content in cigarettes. There are many pharmacologic properties of menthol that may facilitate exposure to tobacco smoke, and it has been suggested that the preference for menthol cigarettes in black smokers accounts for their higher cotinine levels. OBJECTIVE: To assess cigarette per day (cpd)-adjusted cotinine levels in relation to smoking a menthol or nonmenthol cigarette brand among non-Hispanic black and white U.S. adult smokers under natural smoking conditions. METHOD: Serum cotinine concentrations were measured in 1,943 smokers participating in the 2001 to 2006 National Health and Nutrition Examination Surveys (NHANES). The effect of smoking a menthol brand on cpd-adjusted serum cotinine levels in these two populations was modeled by adjusting for sex, age, number of smokers living in the home, body weight, time since last smoked, and FTC-measured nicotine levels. The 8 or 12 digit Universal Product Code (UPC) on the cigarette label was used to determine the cigarette brand and whether it was menthol. RESULTS: Smoking a menthol cigarette brand versus smoking a nonmenthol cigarette brand was not associated (p≥0.05) with mean serum cotinine concentration in either black or white smokers. CONCLUSIONS: The higher levels of cotinine observed in black smokers compared to white smokers is not explained by their higher preference for menthol cigarette brands. IMPACT: Further studies like ours are needed to improve our ability to understand health consequences of future changes in tobacco product design.

BACKGROUND: Prenatal bisphenol A (BPA) exposure may be associated with developmental toxicity, but few studies have examined the variability and predictors of urinary BPA concentrations during pregnancy. OBJECTIVE: Our goal was to estimate the variability and predictors of serial urinary BPA concentrations taken during pregnancy. METHODS: We measured BPA concentrations during pregnancy and at birth in three spot urine samples from 389 women. We calculated the intraclass correlation coefficient (ICC) to assess BPA variability and estimated associations between log10-transformed urinary BPA concentrations and demographic, occupational, dietary, and environmental factors, using mixed models. RESULTS: Geometric mean (GM) creatinine-standardized concentrations (micrograms per gram) were 1.7 (16 weeks), 2.0 (26 weeks), and 2.0 (birth). Creatinine-standardized BPA concentrations exhibited low reproducibility (ICC = 0.11). By occupation, cashiers had the highest BPA concentrations (GM: 2.8 mug/g). Consuming canned vegetables at least once a day was associated with higher BPA concentrations (GM = 2.3 mug/g) compared with those consuming no canned vegetables (GM = 1.6 mug/g). BPA concentrations did not vary by consumption of fresh fruits and vegetables, canned fruit, or store-bought fresh and frozen fish. Urinary high-molecular-weight phthalate and serum tobacco smoke metabolite concentrations were positively associated with BPA concentrations. CONCLUSIONS: These results suggest numerous sources of BPA exposure during pregnancy. Etiological studies may need to measure urinary BPA concentrations more than once during pregnancy and adjust for phthalates and tobacco smoke exposures.

Although clinic-based studies have used biochemical validation to estimate the percentage of pregnant women who deny smoking but are actually smokers, a population-based estimate of nondisclosure of smoking status in US pregnant women has not been calculated. The authors analyzed data from the 1999-2006 National Health and Nutrition Examination Survey and estimated the percentage of 994 pregnant and 3,203 nonpregnant women 20-44 years of age who did not report smoking but had serum cotinine levels that exceeded the defined cut point for active smoking (nondisclosure). Active smoking was defined as self-reporting smoking or having a serum cotinine concentration that exceeded the cut point for active smoking. Overall, 13.0% (95% confidence interval (CI): 8.8, 17.1) of pregnant women and 29.7% (95% CI: 27.3, 32.1) of nonpregnant women were active smokers. Nondisclosure was higher among pregnant active smokers (22.9%, 95% CI: 11.8, 34.6) than among nonpregnant smokers (9.2%, 95% CI: 7.1, 11.2). Among pregnant active smokers, nondisclosure was associated with younger age (20-24 years). Among nonpregnant active smokers, nondisclosure was associated with Mexican-American and non-Hispanic black race/ethnicity. Studies and surveillance systems that rely on self-reported smoking status are subject to underestimation of smoking prevalence, especially among pregnant women, and underreporting may vary by demographic characteristics.

Hair nicotine and cotinine have been proposed as longer-term markers of exposure to secondhand smoke. In this study, we evaluated the rate and extent of nicotine and cotinine deposition into beard hair among six male nonsmokers following a single exposure to 4 mg of nicotine in Nicorette((R)) (nicotine polacrilex) gum. We collected beard hair samples daily for 12 days following exposure and urine samples for 6 days after exposure. Using liquid chromatographic-tandem mass spectrometric analysis, we found that both nicotine and cotinine could be detected in beard samples within 24 h of the exposure and reached a maximum of about 71 pg nicotine and 47 pg cotinine/mg hair, respectively, within 1-2 days, followed by a gradual decline. Compared to beard hair concentrations, nicotine, cotinine, and hydroxycotinine were excreted in urine at much higher levels and also peaked on the day after exposure (mean +/- SD urine cotinine = 300 +/- 183 ng/mL). Our results confirmed that both nicotine and cotinine can be measured in beard hair samples following a single dose of nicotine. However, both the time-course and extent of deposition of these analytes in beard hair in this study differed from the results reported previously from a similar evaluation.

The tobacco-specific carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), a metabolite of the tobacco-specific nitrosamine (TSNA) 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), has been measured in urine samples from all participants aged 6 years and older from the National Health and Nutrition Examination Survey 2007-2008. Participants with a serum cotinine concentration of ≥10 ng/mL were identified as tobacco users, primarily cigarette smokers. Regression models were developed to calculate geometric mean NNAL concentrations adjusted for serum cotinine, urinary creatinine, cigarettes per day, and Federal Trade Commission tar values of the cigarettes smoked. Significant differences were found by gender (p = 0.003) and race/ethnicity (p = 0.022 for non-Hispanic white versus non-Hispanic black smokers), but not by menthol type of the cigarettes. Females and non-Hispanic white smokers had the highest adjusted means for urinary NNAL (353 and 336 pg/mL, respectively). The results from this study demonstrated significant relationships between NNAL concentrations and serum cotinine (p < 0.001) and urine creatinine (p < 0.001). The joint effect of linear and quadratic terms for number of cigarettes smoked per day was also statistically significant (p = 0.001). In addition to addressing current NNK exposure levels, these results will form a baseline for future estimates of tobacco users' exposure to this carcinogen.

Maternal smoking during pregnancy is associated with increased risk of childhood overweight body mass index (BMI). Less is known about the association between prenatal secondhand tobacco smoke (SHS) exposure and childhood BMI. We followed 292 mother-child dyads from early pregnancy to 3 years of age. Prenatal tobacco smoke exposure during pregnancy was quantified using self-report and serum cotinine biomarkers. We used linear mixed models to estimate the association between tobacco smoke exposure and BMI at birth, 4 weeks, and 1, 2 and 3 years. During pregnancy, 15% of women reported SHS exposure and 12% reported active smoking, but 51% of women had cotinine levels consistent with SHS exposure and 10% had cotinine concentrations indicative of active smoking. After adjustment for confounders, children born to active smokers (self-report or serum cotinine) had higher BMI at 2 and 3 years of age, compared with unexposed children. Children born to women with prenatal serum cotinine concentrations indicative of SHS exposure had higher BMI at 2 (mean difference [MD] 0.3 [95% confidence interval -0.1, 0.7]) and 3 (MD 0.4 [0, 0.8]) years compared with unexposed children. Using self-reported prenatal exposure resulted in non-differential exposure misclassification of SHS exposures that attenuated the association between SHS exposure and BMI compared with serum cotinine concentrations. These findings suggest active and secondhand prenatal tobacco smoke exposure may be related to an important public health problem in childhood and later life. In addition, accurate quantification of prenatal secondhand tobacco smoke exposures is essential to obtaining valid estimates.

BACKGROUND: The tobacco-specific nitrosamine NNK (4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone) and its reduction product in the body, NNAL (4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol), are potent pulmonary carcinogens. We have measured total NNAL in the U.S. population of tobacco users and nonsmokers exposed to secondhand smoke (SHS). METHODS: We measured total urinary NNAL (free NNAL plus its glucuronides following hydrolysis) by using a sensitive and specific high-performance liquid chromatography / tandem mass spectrometry method. We calculated the percentage above the LOD, the 50th through 95th percentiles, and in some cases geometric means for groups classified by age, gender and race/ethnicity. RESULTS: Total urinary NNAL was measureable at or above its limit of detection (0.6 pg/mL) in 55% of the study participants, including 44% of nonsmokers. The population distribution of urinary NNAL included smoker and nonsmoker regions similar to the bimodal distribution of serum cotinine, and serum cotinine and total urinary NNAL were strongly correlated (r = 0.92; p<0.001). Among nonsmokers, children had significantly higher concentrations of NNAL than did adults aged ≥20 years (p < 0.001). CONCLUSIONS: Among NHANES participants, total NNAL was found at measurable levels in the urine of 44% of nonsmokers, and in 87.5% of those with substantial SHS exposure (with serum cotinine concentrations of 0.1 to 10 ng/mL). Children aged 6-11 years had the highest NNAL concentrations among all nonsmokers. IMPACT: We describe for the first time the distribution of total urinary NNAL in the entire U.S. population including both smokers and nonsmokers. NNAL was detected in 44% of all nonsmokers.

Urinary 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) and NNAL-glucuronide, which are metabolites of the tobacco-specific nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), are valuable biomarkers of tobacco exposure. In epidemiologic investigations, it is often necessary for samples to be collected and stored for various periods of time prior to analysis, and as a consequence, it is important to determine the stability of these analytes during storage. In this study, two urine pools were stored at six different temperatures ranging from -70 degrees C to 55 degrees C. Free NNAL and NNAL-glucuronide were measured at scheduled time intervals for 28 days, after which samples stored at -20 degrees C and -70 degrees C were further monitored for an additional four years. NNAL-glucuronide was not completely stable at room temperature (ca. 22 degrees C) or above. At those temperatures, a gradual loss of NNAL-glucuronide with concomitant increases in free NNAL was observed. However, both free NNAL and NNAL-glucuronide appeared to be stable for at least four weeks when stored in the refrigerator (4 degrees C) and for at least four years at -20 degrees C and -70 degrees C, respectively. These results suggest that valid NNAL concentrations can be measured in frozen urine samples that have been stored for an extended period of time prior to analysis.

We describe here a hemoglobin adduct assay applied to an analysis of samples from smokers and nonsmokers. The assay includes a sensitive method for quantification of orthotoluidine 2-aminonaphthylene, and 3- and 4-aminobiphenyl hemoglobin adducts in human blood using capillary gas chromatography-tandem mass spectrometry. Basic hydrolysis and derivatization with pentafluoropropionic acid anhydride are followed by programmable temperature vaporization and pseudo on-column capillary gas chromatography with positive electron ionization tandem mass spectrometry analysis. Standard deviation of calibration curves (n = 6) shows that the limits of detection for o-toluidine, 2-aminonaphthylene, and 3- and 4-aminobiphenyl were 0.23, 0.39, 0.30, and 0.24 pg total on-column, respectively. The effective working limit of detection is estimated at approximately 5.22 pg/g Hb and 18.73 pg/g Hb for 4-aminobiphenyl and 2-aminonaphthylene, respectively. In a group that was predominately male and African-American, the level of 4-aminobiphenyl Hb adducts was significantly different between smokers and nonsmokers. Among 93 nonsmokers with serum cotinine concentrations less than 10 ng/mL, the geometric mean (95% CI) concentration of 4-aminobiphenyl was 29.9 pg/g hemoglobin (Hb; 29.4 to 30.4). Conversely, in 100 smokers the 4-aminobiphenyl adducts geometric mean concentration was significantly greater at 73.0 pg/g Hb (72.6 to 73.4). 4-Aminobiphenyl hemoglobin adduct and serum cotinine concentrations were correlated (r = 0.496; p < 0.0001; n = 193). In 15% of smokers, 3-aminobiphenyl was detected at low concentration. Adduct levels of 2-aminonaphthylene and ortho-toluidine were not significantly different between the smoker and nonsmoker participants. Our study shows that 4-aminobiphenyl Hb adducts remain the preferred biomarker for identifying people exposed to aromatic amines from tobacco smoke.

BACKGROUND: Smokers are exposed to significant doses of carcinogens, including tobacco-specific nitrosamines (TSNA). Previous studies have shown significant global differences in the levels of TSNAs in cigarette smoke because of the variation in tobacco blending and curing practices around the world. METHODS: Mouth-level exposure to 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) measured in cigarette butts and urinary concentrations of its major metabolite 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) were examined among 126 daily smokers in four countries over a 24-hour study period. RESULTS: As mouth-level exposure of NNK increased, the urinary NNAL increased even after adjustment for other covariates (beta = 0.46, P = 0.004). The relationship between mouth-level exposure to nicotine and its salivary metabolite, cotinine, was not statistically significant (beta = 0.29, P = 0.057), likely because of the very limited range of differences in mouth-level nicotine exposure in this population. CONCLUSIONS: We have shown a direct association between the 24-hour mouth-level exposure of NNK resulting from cigarette smoking and the concentration of its primary metabolite, NNAL, in the urine of smokers. Internal dose concentrations of urinary NNAL are significantly lower in smokers in countries that have lower TSNA levels in cigarettes such as Canada and Australia in contrast to countries that have high levels of these carcinogens in cigarettes, such as the United States. Impact: Lowering the levels of NNK in the mainstream smoke of cigarettes through the use of specific tobacco types and known curing practices can significantly affect the exposure of smokers to this known carcinogen. (c)2010 AACR.

INTRODUCTION: Body mass index (BMI) and total blood volume are not always considered as variables that affect serum cotinine concentrations. METHOD: We used data from the National Health and Nutrition Examination Survey (NHANES) for the years 1999-2008 and fitted regression models for smokers. In addition to traditionally used covariates like age, race, gender, and average number of cigarettes smoked daily, we used BMI and total blood volume (TBV) as continuous variables to evaluate the impact of these variables on serum cotinine levels. RESULTS: Adjusted serum cotinine levels increased statistically significantly with increase in age (p<0.001). Serum cotinine levels increased statistically significantly (p<0.001) with average number of cigarettes smoked daily. Levels of adjusted serum concentrations from high to low by race/ethnicity were: non-Hispanic blacks, non-Hispanic whites, other race/ethnicity, and Mexican-Americans; and all differences were statistically significant. A model of serum cotinine including BMI without TBV found BMI to be a significant predictor (p<0.001) and similarly a model including TBV without BMI found TBV to be a significant predictor (p<0.001). When BMI and TBV were both included in the model, the significance of BMI changed markedly (p=0.93) with substantive changes in the BMI coefficient and the significance of TBV changed also (p=.024) with small change in the TBV coefficient. DISCUSSION: TBV and BMI are significant predictors of serum cotinine concentrations. TBV or BMI, but not both, should be included in predictive models of serum cotinine concentrations.

Although cigarette smoking remains the most prevalent form of tobacco use in girls and in women of reproductive age globally, use of non-cigarette forms of tobacco is prevalent or gaining in popularity in many parts of the world, especially in low- and middle-income countries. Sparse but growing evidence suggests that the use of some non-cigarette tobacco products during pregnancy increases the risk of adverse pregnancy outcomes. In this paper we review the literature on the prevalence of non-cigarette tobacco product use in pregnant women and in women of reproductive age in high-, middle-, and low-income countries and the evidence that maternal use of these products during pregnancy has adverse health effects. In addition, we communicate findings from an international group of perinatal and tobacco experts that was convened to establish research priorities concerning the use of non-cigarette tobacco products during pregnancy. The working group concluded that attempts to develop a public health response to non-cigarette tobacco use in women are hindered by a lack of data on the epidemiology of use in many parts of the world and by our limited understanding of the type and magnitude of the health effects of these products. We highlight research gaps and provide recommendations for a global research agenda.