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Starting in the 1970s, individuals, businesses and the public have increasingly benefited from policies prohibiting smoking indoors, saving thousands of lives and billions of dollars in healthcare expenditures. Smokefree policies to protect against secondhand smoke exposure, however, do not fully protect the public from the persistent and toxic chemical residues from tobacco smoke (also known as thirdhand smoke) that linger in indoor environments for years after smoking stops. Nor do these policies address the economic costs that individuals, businesses and the public bear in their attempts to remediate this toxic residue. We discuss policy-relevant differences between secondhand smoke and thirdhand smoke exposure: persistent pollutant reservoirs, pollutant transport, routes of exposure, the time gap between initial cause and effect, and remediation and disposal. We examine four policy considerations to better protect the public from involuntary exposure to tobacco smoke pollutants from all sources. We call for (a) redefining smokefree as free of tobacco smoke pollutants from secondhand and thirdhand smoke; (b) eliminating exemptions to comprehensive smoking bans; (c) identifying indoor environments with significant thirdhand smoke reservoirs; and (d) remediating thirdhand smoke. We use the case of California as an example of how secondhand smoke-protective laws may be strengthened to encompass thirdhand smoke protections. The health risks and economic costs of thirdhand smoke require that smokefree policies, environmental protections, real estate and rental disclosure policies, tenant protections, and consumer protection laws be strengthened to ensure that the public is fully protected from and informed about the risks of thirdhand smoke exposure.

IntroductionThis study examined user behavior, e-cigarette dependence, and device characteristics on nicotine intake among users of pod-mod e-cigarettes.Aims and methodsIn 2019-2020, people who use pod-mods in the San Francisco Bay Area completed questionnaires and provided a urine sample for analysis of total nicotine equivalents (TNE). The relationship between TNE and e-cigarette use, e-cigarette brands, e-liquid nicotine strength, e-cigarette dependence, and urine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), as a measure of combustible cigarette exposure, were examined.ResultsOf 100 participants (64% male, 71% in the 18-34 age group, 45% white), 53 used JUUL primarily, 12 used Puff Bar primarily, and 35 used other brands, including Suorin; 48 participants reported current cigarette smoking. Participants most often reported use of e-liquid with 4.5%-6.0% nicotine (68%), fruit (35%), tobacco (28%), and menthol or mint flavors (26%), used e-cigarettes on 25.5 (SD = 6.3) days a month, 10.2 (SD = 14.2) times a day, and 40% used 1-2 pods/cartridges per week. In bivariate analysis, urinary TNE was higher with greater frequency (days used) and intensity (number of pods used) of e-cigarette use, e-cigarette dependence, and combustible cigarette use. In multivariable analysis, days of e-cigarette use in the last 30 days, number of pods used per week, and NNAL levels were significantly associated with TNE. There was no significant impact of e-liquid nicotine strength on TNE.ConclusionsNicotine intake among people who used pod-mod e-cigarettes increased with e-cigarette consumption and e-cigarette dependence, but not with e-liquid nicotine strength. Our findings may inform whether FDA adopts a nicotine standard for e-cigarettes.ImplicationsThe study examined how device and user characteristics influence nicotine intake among pod-mod e-cigarette users. Nicotine intake increased with frequency (days of e-cigarette use in past 30 days) intensity of use (number of pods used per day) and e-cigarette dependence but not with the flavor or nicotine concentration of the e-liquids. Regulation of nicotine concentration of e-liquids is unlikely to influence nicotine exposure among adult experienced pod-mod users.

IntroductionThe aims of this study were to characterize particle size in a thirdhand smoke aerosol and measure the effects of controlled inhalational exposure to thirdhand smoke 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. Thirdhand smoke that is sorbed to surfaces can emit particles back into the air.MethodsSmoke particle size was measured with a scanning mobility particle sizer/condensation particle counter. Using a crossover study design, 18 healthy nonsmokers received a three-hour inhalational exposure to thirdhand smoke and to filtered, conditioned air. Thirdhand smoke 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.ResultsMean smoke particle size increased with aging (171 nm to 265 nm). We found significant increases in urinary cotinine and 3-hydroxycotinine after three hours of exposure to thirdhand smoke and no significant increases in NNAL, interleukin-6, vascular endothelial growth factor or 8-isoprostane.ConclusionsAcute 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 thirdhand smoke exposure screening.ImplicationsThis study shows that a three-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.

Tobacco-specific nitrosamines (TSNAs) are emitted during smoking and form indoors by nitrosation of nicotine. Two of them, N'-nitrosonornicotine (NNN) and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), are human carcinogens with No Significant Risk Levels (NSRLs) of 500 and 14 ng day-1, respectively. Another TSNA, 4-(methylnitrosamino)-4-(3-pyridyl) butanal (NNA), shows genotoxic and mutagenic activity in vitro. Here, we present additional evidence of genotoxicity of NNA, an assessment of TSNA dermal uptake, and predicted exposure risks through different pathways. Dermal uptake was investigated by evaluating the penetration of NNK and nicotine through mice skin. Comparable mouse urine metabolite profiles suggested that both compounds were absorbed and metabolized via similar mechanisms. We then investigated the effects of skin constituents on the reaction of adsorbed nicotine with nitrous acid (epidermal chemistry). Higher TSNA concentrations were formed on cellulose and cotton substrates that were precoated with human skin oils and sweat compared to clean substrates. These results were combined with reported air, dust, and surface concentrations to assess NNK intake. Five different exposure pathways exceeded the NSRL under realistic scenarios, including inhalation, dust ingestion, direct dermal contact, gas-to-skin deposition, and epidermal nitrosation of nicotine. These results illustrate potential long-term health risks for nonsmokers in homes contaminated with thirdhand tobacco smoke.

BackgroundThe Hooked on Nicotine Checklist (HONC) has been used to assess nicotine dependence (loss of autonomy over tobacco) among adolescents. Existing HONC validation studies for non-cigarette products, such as electronic cigarettes (e-cigarettes), have generally not considered biomarkers of nicotine exposure.MethodsWithin a cross-sectional sample of California (USA) high school students (total N = 1396; mean age 15.2 years; 56% female; 54% Hispanic/Latinx), self-reported past 30-day users of any tobacco (including e-cigarettes) completed a modified 10-item HONC questionnaire and provided saliva samples (N = 318 samples, including N = 234 exclusive past 30-day e-cigarette users). Samples were analyzed for cotinine using liquid chromatography-tandem mass spectrometry (lower limit of quantification: 1.0 ng/mL).ResultsAcross four categories of HONC score corresponding to an increasing number of reported dependence symptoms (scores: 0, 1, 2-4, 5-10), the prevalence of quantifiable salivary cotinine increased among past 30-day tobacco users (20%, 21%, 38%, 55%, respectively, P-for-trend

School children may be exposed to secondhand smoke (SHS) either at home, in transit or in social gatherings permitting smoking in their presence. Questionnaires about SHS often underestimate prevalence and extent of exposure. A more accurate tool is the use of biomarkers such as cotinine (COT) and trans-3'-hydrocycotinine (3HC) as biomarkers of SHS exposure, alongside 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), a reduction product in the body of the tobacco-specific nitrosamine, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), both potent carcinogens. We measured urinary COT, 3HC and total NNAL using sensitive and specific high-performance LC-MS/MS methods. The limit of quantification (LOQ) for each assay were 0.05 ng/mL, 0.1 ng/mL and 0.25 pg/mL respectively. The aim of this study was to evaluate the exposure to SHS of school children (9-11 years), from five public schools in the island of Malta, from questionnaire information about smoking at home and verify it by urinary biomarker data of COT, 3HC and NNAL. These biomarkers were measurable in 99.4%, 95.4% and 98.3% of the participating children respectively. From the children reporting smoking at home, 11% had a history of asthma and had COT, 3HC and NNAL geometric mean concentrations double compared to the non-asthmatic group. In has been confirmed that non-smokers exposed to SHS and THS have a higher NNAL/COT ratio than the group identified as smokers according to specific and defined COT threshold levels (despite the fact that a priori, the entire study group was composed of non-smokers). The implication of high measured levels of urinary NNAL in children should be of concern given its potency. A main effects multifactor ANOVA model was developed and the children's house and school locations and the smoking frequency were statistically significant to predict the levels of the three metabolites. For 3HC only, the status of the employment of the mother was also an important predictor.

AimMost of the carcinogenic pollutants coming from tobacco smoking or other combustion processes tend to accumulate in settled house dust (SHD) over time. This study evaluated the load of these pollutants in smokers and non-smokers' houses from relatively fresh SHD collected in five different districts on the island of Malta.MethodsAn improved, efficient extraction method to obtain three fractions from a 200 mg of SHD was developed. It was validated for the analysis of nicotine and polycyclic aromatic hydrocarbons (PAH) by GC-MS/MS and nicotelline and TSNA by LC-MS/MS. Kruskal-Wallis H tests were used to evaluate differences across districts, while a Mann-Whitney U test was used to check differences between smokers and non-smokers' houses. Diagnostic ratios were used to evaluate the carcinogenicity of PAH in SHD in Malta.ResultsFor all analytes, no statistical difference was observed across different districts, but, in smokers' houses, 97.9% of the total concentration of all target analytes found in SHD is nicotine, 0.1% is TSNA, and 2.0% is PAH. In non-smokers' houses, nicotine represents 16.8% of the load, while 0.4% and 82.8% are TSNA and PAH, respectively. The carcinogenicity of the PAH mixture in Maltese SHD, expressed as the mean benzo(a)pyrene equivalent (BaPeq) is 371 ng/g.ConclusionIndoor activities, ventilation practices, and infiltration of outdoor pollutants contribute to a complex SHD composition. Although the BaPeq is on the lower end of carcinogenicity, the effects of a mixture including tobacco-related potent carcinogens in SHD are largely unknown. In view of indoor, continuous exposure to SHD through several pathways, further research is warranted.

Biomarkers for the use of electronic nicotine delivery systems (ENDS) are desirable for studies of the health effects of electronic cigarettes and related devices. However, the aerosols inhaled from these devices do not contain substances that are unique to this class of products, i.e., substances that are not present in cigarette smoke or those that do not have common environmental or dietary sources. Consequently, identifying selective biomarkers for ENDS use remains a challenge. If co-use of conventional tobacco products can be definitively ruled out, then nicotine and its metabolites are suitable for assessing exposure. Self-reports from questionnaires are often used to obtain information on product use. But self-reports may not always be accurate, and are not amenable to obtaining quantitative information on exposure. An alternative approach is to use selective biomarkers for conventional tobacco products to definitively rule out their use. In this article, we describe two new LC-MS/MS methods for the minor tobacco alkaloids anabasine, anatabine, nicotelline, anatalline, and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), a tobacco-specific nitrosamine metabolite, all biomarkers that are selective for the use of conventional tobacco products. Applications of these biomarkers in studies of ENDS use and dual use of ENDS and conventional tobacco products are also discussed.

ObjectivesTo investigate if the nicotine metabolite ratio (NMR, the ratio of nicotine metabolites 3'-hydroxycotinine/cotinine) is a reliable phenotypic biomarker for nicotine clearance across races, and as a function of differences in the rate of nicotine, cotinine and 3'-hydroxycotinine glucuronidation and UGT genotypes.MethodsParticipants [Caucasians (Whites), African Americans (Blacks) and Asian-Americans (Asians)] received an oral solution of deuterium-labeled nicotine and its metabolite cotinine. Plasma and saliva concentrations of nicotine and cotinine were used to determine oral clearances. Rates of glucuronidation were assessed from urine glucuronide/parent ratios, and UGT2B10 and UGT2B17 genotypes from DNA.ResultsAmong the 227 participants, 96 (42%) were White, 67 (30%) Asian and 64 (28%) Black. Compared to the other two races, Whites had higher nicotine and cotinine total oral clearance, Blacks had lower nicotine and cotinine glucuronidation rates and Asians had lower 3'-hydroxycotinine glucuronidation rates. A strong positive correlation (correlations coefficients 0.77-0.84; P

IntroductionMicrobiome differences have been found in adults who smoke cigarettes compared to non-smoking adults, but the impact of thirdhand smoke (THS; post-combustion tobacco residue) on hospitalized infants' rapidly developing gut microbiomes is unexplored. Our aim was to explore gut microbiome differences in infants admitted to a neonatal ICU (NICU) with varying THS-related exposure.MethodsForty-three mother-infant dyads (household member[s] smoke cigarettes, n = 32; no household smoking, n = 11) consented to a carbon monoxide-breath sample, bedside furniture nicotine wipes, infant-urine samples (for cotinine [nicotine's primary metabolite] assays), and stool collection (for 16S rRNA V4 gene sequencing). Negative binomial regression modeled relative abundances of 8 bacterial genera with THS exposure-related variables (i.e., household cigarette use, surface nicotine, and infant urine cotinine), controlling for gestational age, postnatal age, antibiotic use, and breastmilk feeding. Microbiome-diversity outcomes were modeled similarly. Bayesian posterior probabilities (PP) ≥75.0% were considered meaningful.ResultsA majority of infants (78%) were born pre-term. Infants from non-smoking homes and/or with lower NICU-furniture surface nicotine had greater microbiome alpha-diversity compared to infants from smoking households (PP ≥ 75.0%). Associations (with PP ≥ 75.0%) of selected bacterial genera with urine cotinine, surface nicotine, and/or household cigarette use were evidenced for 7 (of 8) modeled genera. For example, lower Bifidobacterium relative abundance associated with greater furniture nicotine (IRR

Second Hand Smoke (SHS) has always been primarily linked with indoor pollution. To date nicotine was the favoured marker for SHS alongside measurements of particulate matter (PM) levels. As nicotine is mainly found in the gas-phase and reactive in the outdoor environment it is not ideal as a marker for the SHS-driven particulate component in PM. Nicotelline, a minor tobacco alkaloid that is stable, found almost exclusively in the particle phase and easy to quantify even at low concentrations, is being proposed as a better marker. It is the first study using bisulfate-treated quartz fiber filters to show that airborne nicotine (gas+particle phase) is directly proportional to airborne nicotelline in countries that have different climates. The analytical method developed has been validated to show that the use of untreated filters is suitable for the quantification of nicotelline even at low concentrations. Although nicotelline exhibits a seasonal and geographical variation, this is the first comprehensive study which demonstrates the ubiquitous presence of nicotelline in PM from outdoor air samples collected in the USA (0.1-285.6 pgm-3), UK (2.3-9.1 pgm-3), Hong Kong (3.8-109.3 pgm-3) and Malta (4.2-280.8 pgm-3). From the nicotelline apportionment factor of 1589 ng/mg of tobacco smoke PM we estimate the fraction of outdoor airborne PM derived from SHS to be in the range of 0.03-0.08%. While it is unlikely for tobacco smoke-related toxics in outdoor PM to be considered a major health hazard, in heavily polluted microenvironments this marker would be useful in tracing the presence of SHS and emerging Third Hand Smoke components that form or are found in airborne and settled PM that could induce serious health effects.

Thirdhand smoke (THS) is an environmental contaminant that may cause adverse health effects in smokers and nonsmokers. Currently, time-consuming analytical methods are necessary to assess chemicals in THS repositories, like upholstered furniture and clothing. Our goal was to develop a rapid, accessible method that can be used to measure THS contamination in common household fabrics and to evaluate remediation. Cotton, terry cloth, polyester, and wool were exposed to THS for various times in a controlled laboratory environment and then extracted in various media at room temperature or 60 °C to develop an autofluorescent method to quantify THS. Concentrations of nicotine and related alkaloids in the extracts were determined using liquid chromatography-tandem mass spectrometry (LC-MS/MS) and high-performance liquid chromatography (HPLC). The autofluorescence of extracts was proportional to the time and amount of THS exposure received by cotton and terry cloth. Extracts of polyester and wool did not show autofluorescence unless heat was applied during extraction. Nicotine, nicotine alkaloids, and TSNA concentrations were higher in THS extracts from cotton and terry cloth than extracts of polyester and wool carpet, in agreement with the autofluorescence data. For fabrics spiked with 10 mg of nicotine, extraction efficiency was much higher from terry cloth (7 mg) than polyester (0.11 mg). In high relative humidity, nicotine recovery from both cotton and polyester was 80% (~8 mg). Our results provide a simple, rapid method to assess THS contaminants in household fabrics and further show that THS extraction is influenced by fabric type, heat, and humidity. Thus, remediation of THS environments may need to vary depending on the fabric reservoirs being treated. Understanding the dynamics of THS in fabrics can help set up appropriate remediation policies to protect humans from exposure.

ObjectiveIt is unclear why Black smokers in the United States have elevated risk of some tobacco-related diseases compared to White smokers. One possible causal mechanism is differential intake of tobacco toxicants, but results across studies are inconsistent. Thus, we examined racial differences in biomarkers of toxic volatile organic compounds (VOCs) present in tobacco smoke.MethodWe analyzed baseline data collected from 182 Black and 184 White adult smokers who participated in a randomized clinical trial in 2013-2014 at 10 sites across the United States. We examined differences in urinary levels of ten VOC metabolites, total nicotine equivalents (TNE), and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), controlling for covariates such as cigarettes per day (CPD), as well as differences in VOCs per TNE to assess the extent to which tobacco exposure, and not metabolic factors, accounted for racial differences.ResultsConcentration of metabolites of acrolein, acrylonitrile, ethylene oxide, and methylating agents were significantly higher in Blacks compared to Whites when controlled for covariates. Other than the metabolite of methylating agents, VOCs per TNE did not differ between Blacks and Whites. Concentrations of TNE/CPD and VOCs/CPD were significantly higher in Blacks. Menthol did not contribute to racial differences in VOC levels.ConclusionsFor a given level of CPD, Black smokers likely take in higher levels of acrolein, acrylonitrile, and ethylene oxide than White smokers. Our findings are consistent with Blacks taking in more nicotine and toxicants per cigarette smoked, which may explain their elevated disease risk relative to other racial groups.

Background Cardiovascular safety is an important consideration regarding the benefits versus risks of electronic cigarette use (EC) for public health. The single-use cardiovascular effects of EC have been well studied but may not reflect effects of ad libitum use throughout the day. We aimed to compare the circadian hemodynamic effects as well as 24-hour biomarkers of oxidative stress, and platelet aggregation and inflammation, with ad libitum cigarette smoking (CS) versus EC versus no tobacco product use. Methods and Results Thirty-six healthy dual CS and EC users participated in a crossover study in a confined research setting. Circadian heart rate, blood pressure and plasma nicotine levels, 24-hour urinary catecholamines, 8-isoprostane and 11-dehydro-thromboxane B2, and plasma interleukin-6 and interleukin-8 were compared in CS, EC, and no nicotine conditions. Over 24 hours, and during daytime, heart rate and blood pressure were higher in CS and EC compared with no tobacco product conditions (P

IntroductionApproximately the same percentage of male high school students in the United States currently uses conventional smokeless tobacco as smokes cigarettes, resulting in toxin exposure.MethodsThis study assessed tobacco product use (smokeless, combustible, and electronic cigarettes) and nicotine and carcinogen exposures in a sample of 594 male rural high school baseball players-a population traditionally at risk for smokeless tobacco use. Salivary specimens were assayed for cotinine (a biomarker of nicotine exposure) and urine specimens for 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL, a biomarker of the carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) using liquid chromatography-tandem mass spectrometry.ResultsThe prevalence of past 30-day use of any tobacco product was 29%. Past 7-day smokeless tobacco use (prevalence: 13%) was associated with the highest levels of cotinine and NNAL observed in the sample, whether smokeless tobacco was used exclusively (geometric means: cotinine 11.1 ng/mL; NNAL 31.9 pg/mg-creatinine) or in combination with combustible products (geometric means: cotinine 31.6 ng/mL; NNAL 50.0 pg/mg creatinine). Cotinine and NNAL levels were incrementally higher in each increasing category of smokeless tobacco use frequency. However, observed levels were lower than previously reported for adults, likely reflecting less smokeless use per day among adolescents.ConclusionsBased on these biomarker observations, adolescents who use conventional smokeless tobacco products are exposed to substantial levels of nicotine and NNK. Although exposed to lower levels than adult smokeless users, the findings are concerning given the young age of the sample and tendency for smokeless tobacco users to increase use intensity over time.ImplicationsThis study demonstrates that adolescents using smokeless tobacco are exposed to levels of nicotine and NNK that increase with use frequency and that exceed exposures among peers using other tobacco products. Youth smokeless tobacco use in the United States has not declined along with youth smoking prevalence, giving greater importance to this health concern. To reduce youth (and adult) exposures, needed actions include effective smokeless tobacco use prevention, potentially in combination with reducing the levels of harmful and potentially harmful chemicals in smokeless tobacco products currently popular among adolescents.

IntroductionDual use of electronic cigarettes (e-cigarettes) and combustible cigarettes is a major public health issue. It is generally accepted that exclusive e-cigarette use is less harmful than exclusive combustible cigarette use, but most e-cigarette users continue to smoke combustible cigarettes as well. To what extent the use of e-cigarettes reduces harm in people who continue to smoke combustible cigarettes has been debated. The aim of this study was to explore the utility of biomarkers as measures of dual use.MethodsIn two human studies of participants who used e-cigarettes only or both combustible cigarettes and e-cigarettes, we measured urine concentrations of the metabolites of nicotine (total nicotine equivalents) as well as two biomarkers of tobacco exposure: 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), a tobacco-specific carcinogen metabolite, and nicotelline, a tobacco alkaloid not found in significant concentrations in e-cigarette products.ResultsThe presence of nicotine metabolites indicates either e-cigarette or combustible cigarette use. Nicotelline (half-life of 2-3 hours) indicates recent combustible cigarette use and NNAL (half-life of 10 days or more), indicates combustible cigarette use occurring within several weeks prior to sample collection.ConclusionsNicotelline and NNAL are useful biomarkers for combustible tobacco use in users e-cigarettes. The application of these biomarkers provides a tool to help assess whether, or to what extent, dual use of e-cigarettes and combustible cigarettes reduces harm compared to sole use of combustible cigarettes. These biomarkers can also verify exclusive use of e-cigarettes over short (24 hour) or long (several week) time periods.ImplicationsTo what extent dual use of e-cigarettes and combustible cigarettes reduce harm compared to smoking combustible cigarettes only is of considerable public health interest. We show that the levels of the minor tobacco alkaloid nicotelline and the nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) are extremely low in electronic cigarette fluids. The urine biomarkers nicotelline and the NNK metabolite 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) are indicative of cigarette smoking and can be used to assess recent and past smoking in dual users.

BackgroundThe 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.MethodsThe 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.ResultsBiochemical 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).ConclusionsRecommendations 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.ImplicationsBiochemical 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.

IntroductionMany electronic cigarette manufacturers have begun offering liquids containing "nicotine salts," which are formed when an acid is mixed in a solution with free-base nicotine. Type of salt could play a significant role in the abuse liability of electronic cigarette liquids. As a first step to understanding nicotine salts, this study sought to identify the types of acids present in 23 commercially available electronic cigarette liquids.Aims and methodsTwenty-three electronic cigarette liquids advertised as containing nicotine salts were purchased for analysis. These liquids were tested for the presence of 11 different organic acids that were deemed likely to be used in a nicotine salt formulation. Liquids were analyzed using a combination of liquid chromatography-mass spectrometry and gas chromatography-mass spectrometry methods, then compared to authentic acid standards for identification.ResultsSix of the 11 possible acids were identified in the liquids, from most to least common: lactic, benzoic, levulinic, salicyclic, malic, and tartaric acid. Acid(s) could not be identified in one of the liquids. Though most liquids contained only one type, three of the liquids contained multiple acids.ConclusionsThese data demonstrate that several types of salts/acids are currently being used in electronic cigarette liquids. The type and concentration of salt(s) used in these liquids may differentially alter sensations in the throat and upper airway, and overall pharmacology of the aerosols by altering liquid pH and from flavor and sensory characteristics of the acids themselves.ImplicationsThis study demonstrates that at least six different types of acids are being used to create the nicotine salts in electronic cigarette liquids, with the acids lactic, benzoic, and levulinic being the most frequently identified. Identification of these acids can serve as the foundation for future research to determine if type of nicotine salt alters pharmacological and toxicological effects of electronic cigarettes.

ObjectiveIn an urban adolescent population, we evaluated sources of exposure to secondhand smoke exposure (SHS), examined differences in exposure by race/ethnicity, age and sex, and determined the relationship between exposure source(s) and the biomarkers cotinine and NNAL.MethodsParticipants were recruited from a public hospital-based outpatient clinic in San Francisco, CA, USA.ResultsOf a sample of N = 298 adolescents screened, 235 were biologically confirmed to be exposed to tobacco smoke. Of those, N = 16 were active smokers and N = 219 were exposed to SHS; 91 (39%) were heavily SHS exposed (median cotinine = 0.76 ng/mL) and 128 (54%) had light SHS exposure (median cotinine = 0.11 ng/mL). Within those SHS exposed, the most common source of exposure was in a public area. No significant racial/ethnic differences were found, although African American adolescents were more likely to live in a home that allowed smoking. Older adolescents were more likely to be exposed across several difference sources, and females more likely to be exposed in a car and in public areas. Past 7-day exposure in the home, in a car, and current blunt use were significantly related to biomarkers of exposure.ConclusionsUrban adolescents are exposed to SHS across a variety of sources. Although exposure in a public area is most common, exposure in the home and in cars significantly influences tobacco biomarker levels. Interventions to reduce exposure would have the greatest impact in this population if they focused on reducing exposure in the home and in cars. History of blunt use is a strong determinant of tobacco exposure.

AimTo describe systemic nicotine exposure and subjective effects of electronic cigarettes (e-cigarettes) in people who use both e-cigarettes and cigarettes (dual users), including within-subject comparisons of e-cigarette and cigarette use.DesignTwo-arm, counterbalanced cross-over study. Participants used their usual brand of e-cigarette or cigarette during a standardized session in a 2-week study.SettingHospital research ward, San Francisco, CA, USA.ParticipantsThirty-six healthy (eight women, 28 men) participants.MeasurementsPlasma nicotine was analyzed by gas chromatography-tandem mass spectrometry; nicotine withdrawal, urge to smoke and vape, affective states, craving, satisfaction and psychological reward were measured by standardized questionnaires.FindingsCompared with cigarettes, average maximum plasma nicotine concentration (Cmax ) was lower with e-cigarettes [6.1 ± 5.5 ng/ml, mean ± standard deviation (SD) versus 20.2 ± 11.1 ng/ml, P

IntroductionAccurate measurement of nicotine exposure from cigarette smoke is important in studying disease risk and level of dependence. Urine total nicotine equivalents, the molar sum of nicotine and six metabolites (NE7), accounts for more than 90% of a nicotine dose and is independent of individual metabolic differences. However, measuring NE7 is technically difficult and costly. We compared NE7, the gold standard of nicotine intake, with different combinations of fewer urinary nicotine metabolites. We also examined the impact of individual differences in nicotine metabolic rate, sex, and race on strength of association with NE7.MethodsUrine samples from 796 daily smokers, who participated across five clinical studies, were assayed for nicotine and/or metabolites. Associations with NE7 were assessed by regression and Bland-Altman analyses.ResultsOverall, the molar sum of urine [cotinine + 3'-hydroxycotinine (3HC)] (NE2) and [nicotine + 
cotinine + 3HC] (NE3) were strongly correlated with NE7 (r = .97 and .99, respectively). However, in slow metabolizers NE2 was less predictive of NE7, whereas NE3 was equally robust. Urine total cotinine was also strongly correlated with NE7 (r = .87).ConclusionsUrine NE3 is a robust biomarker of daily nicotine intake, independently of individual metabolic differences, whereas NE2 is less accurate in slow metabolizers. Our findings inform the selection of more rigorous and cost-effective measures to assess nicotine exposure in tobacco research studies.ImplicationsThe molar sum of urine total nicotine, cotinine and 3HC (NE3) is a robust biomarker of daily nicotine intake, independently of individual metabolic differences, and performs as well as measuring seven nicotine metabolites (NE7). The sum of cotinine and 3HC (NE2) is less accurate in slow metabolizers. Our findings inform the selection of more rigorous and cost-effective measures to assess nicotine exposure in tobacco research studies.

Comparisons of systemic exposure to toxicants during monitored cigarette smoking, electronic cigarette (e-cigarette) use, and abstention are needed to enhance our understanding of the risks of e-cigarette use (vaping). In a cross-over study, we measured 10 mercapturic acid metabolites of volatile organic compounds (VOCs) in 24-hour urine samples collected from 36 dual users (8 women) of e-cigarettes and cigarettes during 2 days of ad libitum vaping or cigarette-only use, and 2 days of enforced abstention. Concentrations of VOC metabolites were higher during smoking compared with vaping, except for the methylating agents' metabolite. The fold-difference in concentrations when smoking relative to vaping ranged from 1.31 (1.06-1.61; geometric mean, 95% confidence interval; 1,3-butadiene) to 7.09 (5.88-8.54; acrylonitrile). Metabolites of acrylamide [fold difference of 1.21 (1.03-1.43)] and benzene [1.46 (1.13-1.90)] were higher during vaping compared with abstention. The 1,3-butadiene and propylene oxide metabolites were higher in variable-power tank users compared with users of cig-a-likes. E-cigarettes expose users to lower levels of toxic VOCs compared with cigarette smoking, supporting their harm reduction potential among smokers. However, some e-cigarettes expose users to VOCs such as acrylamide, benzene, and propylene oxide, and may pose health risks to nonsmoking users. The results of our study will inform regulators in assessing e-cigarettes with respect to the balance between its potential harm reduction for adult smokers and risk to nonsmoking users.

BackgroundAssessing the prevalence and level of exposure (dose) of tobacco and marijuana use is important in studies of harm from use of these substances. We used biochemical analysis of urine to quantitatively assess exposure to nicotine and delta 9-tetrahydrocannabinol (THC) in adolescents receiving medical care in a public hospital METHODS: Participants were 686 adolescents between 12 and 21 years old seen at Zuckerberg San Francisco General Hospital between 2012 and 2014. Urine samples were assayed using high sensitivity liquid chromatographic assays for cotinine, a major metabolite of nicotine, and 11-nor-9-carboxy-delta 9-THC (THC-COOH), a major metabolite of THC. A commonly used immunoassay screen for THC-COOH was also performed.ResultsThe THC-COOH immunoassay substantially underestimated THC exposure, as measured with the high sensitivity assay. THC use was detected in 25% of participants, with higher prevalence with increasing age and in non-Hispanic blacks. Active tobacco smokers had an 80% prevalence of THC use (odds ratio for cigarette smoking predicting THC use 13.2). Urine cotinine and THC-COOH were significantly correlated (r = 0.60).ConclusionsThe use of a high sensitivity chromatographic urine assay provides a much more complete picture of adolescent tobacco use compared to a commonly used immunoassay. The immunoassay provides high specificity but moderate sensitivity. We confirm high concordance of tobacco and marijuana use and the high predictive value of cigarette smoking in predicting marijuana use, and provide novel data on the quantitative correlation between level of exposure to nicotine and THC. Quantitative screening of nicotine and THC exposure may enhance our understanding of addiction and harm from single and dual product use.

BackgroundBlacks bear a disproportionate burden of smoking-related diseases and experience greater difficulty quitting smoking than Whites. Nicotine has a high affinity for melanin, and it has been hypothesized that melanin levels might influence nicotine pharmacokinetics and enhance dependence. The aim of this study was to evaluate the hypothesis that melanin affects nicotine disposition kinetics in humans.MethodsForty-four Black participants were administered intravenous infusions of deuterium-labeled nicotine and cotinine. Plasma concentrations of nicotine and cotinine were measured, and pharmacokinetic parameters were estimated. The constitutive and facultative melanin indexes were measured using a dermaspectrophotometer.ResultsThe median constitutive melanin index was 60.7 (32.8-134.7) and the median facultative melanin index 68.1 (38.6-127.1). The mean (±SD) nicotine elimination half-life was 136 min (±33.5), clearance was 1237 mL/min (±331), and Vss was 204 L (±66), or 2.6 L/kg (±0.7). No evidence of significant differences was found in nicotine pharmacokinetic parameters by comparing participants in different melanin index quartiles (outliers with very high melanin index had similar pharmacokinetic values to others). Differences were not statistically significant when adjusted for age, BMI, sex and CYP2A6 genotype or the nicotine metabolite ratio (NMR), and no evidence of significant correlations were found between melanin (facultative or constitutive) and the pharmacokinetic parameters of nicotine or cotinine or tobacco dependence measures.ConclusionsBased on our finding in this group of Black smokers, we could not confirm the hypothesis that melanin significantly affects nicotine disposition kinetics or measures of tobacco dependence.

BackgroundElectronic cigarettes (ECs) are nicotine delivery devices that produce aerosol without combustion of tobacco; therefore, they do not produce sidestream smoke. Nevertheless, many users exhale large clouds of aerosol that can result in passive exposure of non-users. Analogous to thirdhand cigarette smoke, the exhaled aerosol also settles on indoor surfaces where it can produce a residue. We refer to this residue as EC exhaled aerosol residue (ECEAR). Our objective was to determine if exhaled EC aerosol transferred from a vape shop in a multiple-tenant retail building, where it was produced, to a nearby business (field site) where it could deposit as ECEAR.MethodsWe examined the build-up of ECEAR in commonly used materials (cotton towel and paper towels) placed inside the field site across from the vape shop. Materials were subjected to short-term (days) and long-term (months) exposures. Nicotine, other alkaloids and tobacco-specific nitrosamines (TSNAs) were identified and quantified in controls and field site samples using analytical chemical techniques.ResultsNicotine and other alkaloids were detected after 1 day of exposure in the field site, and these chemicals generally increased as exposure times increased. TSNAs, which have been linked to carcinogenesis, were also detected in short-term and long-term exposed samples from the field site.ConclusionsIn a multiple-tenant retail building, chemicals in EC aerosol travelled from a vape shop into an adjacent business where they deposited forming ECEAR. Regulatory agencies and tenants occupying such buildings should be aware of this potential environmental hazard.

ObjectiveTo determine whether early vascular aging may be present in flight attendants with remote in-cabin secondhand smoke (SHS) exposure.MethodsTwenty-six flight attendants with a history of in-cabin SHS exposure prior to the airline smoking bans were recruited. Pulse wave analysis, peripheral arterial tonometry, and brachial artery reactivity testing evaluated their arterial compliance and endothelial function.ResultsFlight attendants with remote in-cabin SHS exposure have normal blood pressure, pulse wave velocity, and reactive hyperemia index, but abnormal pulse pressure, augmentation index, flow-mediated dilation, and hyperemic mean flow ratio.ConclusionThese preliminary findings suggest that flight attendants with remote in-cabin SHS exposure have preclinical signs of accelerated vascular aging and raise new questions about the relationship between remote SHS exposure and vascular health.

BackgroundElectronic cigarette (EC) users may exhale large clouds of aerosol that can settle on indoor surfaces forming ECEAR (EC exhaled aerosol residue). Little is known about the chemical composition or buildup of this residue.ObjectiveOur objective was to identify and quantify ECEAR chemicals in two field sites: an EC user's living room and a multi-user EC vape shop.MethodsWe examined the buildup of ECEAR in commonly used materials (cotton, polyester, or terrycloth towel) placed inside the field sites. Materials were subjected to different lengths of exposure. Nicotine, nicotine alkaloids, and tobacco-specific nitrosamines (TSNAs) were identified and quantified in unexposed controls and field site samples using analytical chemical techniques.ResultsNicotine and nicotine alkaloids were detected in materials inside the EC user's living room. Concentrations of ECEAR chemicals remained relatively constant over the first 5 months, suggesting some removal of the chemicals by air flow in the room approximating a steady state. ECEAR chemicals were detected in materials inside the vape shop after 6 h of exposure and levels continually increased over a month. By 1 month, the nicotine in the vape shop was 60 times higher than in the EC user's living room. ECEAR chemical concentrations varied in different locations in the vape shop. Control fabrics had either no detectable or very low concentrations of chemicals.ConclusionsIn both field sites, chemicals from exhaled EC aerosols were deposited on indoor surfaces and accumulated over time forming ECEAR. Non-smokers, EC users, and employees of vape shops should be aware of this potential environmental hazard.

BackgroundSecondhand smoke (SHS) in US casinos is common, but little is known about the residue of tobacco smoke pollutants left behind in dust and on surfaces, commonly referred to as thirdhand smoke (THS). We examined SHS and THS pollution and exposure before and during a casino smoking ban and after smoking resumed.MethodsA casino was visited nine times over a 15-month period to collect dust, surface and air samples in eight locations. Finger wipe and urine samples were collected from non-smoking confederates before and after a 4-hour casino visit. Samples were analysed for markers of SHS and THS pollution and exposure.ResultsExceptionally high levels of THS were found in dust and on surfaces. Although the smoking ban led to immediate improvements in air quality, surface nicotine levels were unchanged and remained very high for the first month of the smoking ban. Surface nicotine decreased by 90% after 1 month (P

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 that are currently active in this area. Each laboratory assayed blind replicates of seven human urine pools at various concentrations on three separate days. The samples included five pools blended from smoker and nonsmoker urine sources, and two 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 < + 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. Cancer Epidemiol Biomarkers Prev; 27(9); 1083-90. ©2018 AACR.

Background: Many adolescents are exposed to tobacco smoke, from either active smoking (CS) or secondhand smoke (SHS) exposure. Tobacco-specific biomarkers of exposure include cotinine (detects use in past 2-4 days) and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL; detects use for a month or longer). NNAL is expected to detect more intermittent tobacco exposure. We compared NNAL and cotinine as biomarkers of exposure to tobacco in urban adolescents and determined the optimal NNAL cutoff point to distinguish CS from SHS exposure.Methods: Surplus urine samples, collected from 466 adolescents attending pediatric well or urgent care visits at Zuckerberg San Francisco General Hospital in 2013 to 2014, were assayed for cotinine and NNAL.Results: Ninety-four percent of adolescents had measurable levels of NNAL compared with 87% for cotinine. The optimal NNAL cutoff point to distinguish CS from SHS was 9.6 pg/mL by latent class or 14.4 pg/mL by receiver-operating characteristic analysis. Cotinine and NNAL were strongly correlated, but the correlation slopes differed for active versus SHS-exposed adolescents. Among nonsmokers, NNAL levels were significantly higher in African American (median, 3.3 pg/mL) compared with other groups (0.9-1.9 pg/mL), suggesting greater exposure to SHS.Conclusions: Urine NNAL screening finds a large majority (94%) of urban adolescents are exposed to tobacco. African Americans are exposed to higher levels of SHS than other ethnic/racial groups.Impact: SHS is associated with significant medical morbidity in adolescents. Routine biochemical screening with NNAL or cotinine detects high prevalence of SHS exposure and should be considered as a tool to reduce SHS exposure in high-risk populations. Cancer Epidemiol Biomarkers Prev; 27(3); 254-61. ©2018 AACR.

Exposure to thirdhand smoke (THS) is a recently described health concern that arises in many indoor environments. However, the carcinogenic potential of THS, a critical consideration in risk assessment, remains untested. Here we investigated the effects of short-term early exposure to THS on lung carcinogenesis in A/J mice. Forty weeks after THS exposure from 4 to 7 weeks of age, the mice had increased incidence of lung adenocarcinoma, tumor size and, multiplicity, compared with controls. In vitro studies using cultured human lung cancer cells showed that THS exposure induced DNA double-strand breaks and increased cell proliferation and colony formation. RNA sequencing analysis revealed that THS exposure induced endoplasmic reticulum stress and activated p53 signaling. Activation of the p53 pathway was confirmed by an increase in its targets p21 and BAX. These data indicate that early exposure to THS is associated with increased lung cancer risk.

IntroductionCharacterization of aerosols generated by electronic cigarettes (e-cigarettes) is one method used to evaluate the safety of e-cigarettes. While some researchers have modified smoking machines for e-cigarette aerosol generation, these machines are either not readily available, not automated for e-cigarette testing or have not been adequately described. The objective of this study was to build an e-cigarette vaping machine that can be used to test, under standard conditions, e-liquid aerosolization and nicotine and toxicant delivery.MethodsThe vaping machine was assembled from commercially available parts, including a puff controller, vacuum pump, power supply, switch to control current flow to the atomizer, three-way value to direct air flow to the atomizer, and three gas dispersion tubes for aerosol trapping. To validate and illustrate its use, the variation in aerosol generation was assessed within and between KangerTech Mini ProTank 3 clearomizers, and the effect of voltage on aerosolization and toxic aldehyde generation were assessed.ResultsWhen using one ProTank 3 clearomizer and different e-liquid flavors, the coefficient of variation (CV) of aerosol generated ranged between 11.5% and 19.3%. The variation in aerosol generated between ProTank 3 clearomizers with different e-liquid flavors and voltage settings ranged between 8.3% and 16.3% CV. Aerosol generation increased linearly at 3-6V across e-liquids and clearomizer brands. Acetaldehyde, acrolein, and formaldehyde generation increased markedly at voltages at or above 5V.ConclusionThe vaping machine that we describe reproducibly aerosolizes e-liquids from e-cigarette atomizers under controlled conditions and is useful for testing of nicotine and toxicant delivery.ImplicationsThis study describes an electronic cigarette vaping machine that was assembled from commercially available parts. The vaping machine can be replicated by researchers and used under standard conditions to generate e-cigarette aerosols and characterize nicotine and toxicant delivery.

Tobacco smoking and exposure to tobacco secondhand smoke (SHS) can cause lung cancer. We determined uptake of NNK (4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone), a tobacco specific potent pulmonary carcinogen, in hookah smokers and non-smokers exposed to hookah tobacco SHS. We analyzed data from a community-based convenience sample of 201 of adult (aged ≥18 years) exclusive hookah smokers (n = 99) and non-smokers (n = 102) residing in San Diego County, California. Participants spent an average of three consecutive hours indoors, in hookah lounges or private homes, where hookah tobacco was smoked exclusively. Total NNAL [the sum of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) and its glucuronides], the major metabolites of NNK, were quantified in spot urine samples provided the morning of and the morning after attending a hookah event. Among hookah smokers urinary NNAL increased significantly (p

IntroductionRoutine biochemical assessment of tobacco smoke exposure could lead to more effective interventions to reduce or prevent secondhand smoke (SHS)-related disease in adolescents. Our aim was to determine using urine cotinine (major nicotine metabolite) measurement the prevalence of tobacco smoke exposure among adolescents receiving outpatient care at an urban public hospital.MethodsSurplus urine was collected in 466 adolescents attending pediatric or urgent care clinics at Zuckerberg San Francisco General Hospital, serving families with lower levels of income and education, in 2013-2014. The majority were Hispanic or African American. Urine cotinine cut points of 0.05 to 0.25 ng/ml, 0.25 to 30 ng/ml, and 30 ng/ml were used to classify subjects as light SHS or thirdhand smoke exposed, SHS or light/intermittent active users, and active tobacco users, respectively.ResultsAmong subjects 87% were exposed, including 12% active smoking, 46% SHS and 30% lightly exposed. The SHS exposed group adjusted geometric mean cotinine values were significantly higher in African Americans (1.48 ng/ml) compared to other groups (0.56-1.13 ng/ml).ConclusionsIn a city with a low smoking prevalence (12%), a large majority (87%) of adolescents seen in a public hospital clinic are exposed to tobacco. This is much higher than reported in national epidemiological studies of adolescents, which used a plasma biomarker. Since SHS is associated with significant respiratory diseases and parents and adolescents underreport exposure to SHS, routine biochemical screening should be considered as a tool to reduce SHS exposure. The clinical significance of light exposure needs to be investigated.ImplicationsUrine biomarker screening found that a large majority (87%) of adolescents treated in an urban public hospital are exposed to tobacco. Since SHS is associated with significant respiratory diseases and parents and adolescents underreport exposure to SHS, routine biochemical screening should be considered as a tool to reduce SHS exposure.

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 use 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 pipes, and cigarillos. Rigorously validated biomarkers specific to these new products have not yet been identified. Here, we discuss the strengths and limitations of current approaches, including whether they provide reliable exposure estimates for new and emerging products. 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 and avoid common experimental errors. With the identification of the capacity gaps in biomarker research on new and emerging tobacco products, we hope to provide researchers, policymakers, and funding agencies with a clear action plan for conducting and promoting research on the patterns of use and health effects of these products.

ObjectivesTo describe the effect of e-liquid flavors on nicotine intake and pharmacology of e-cigarettes.Methods11 males and 3 females participated in a 3-day inpatient crossover study with strawberry, tobacco, and their usual flavor e-liquid. Nicotine levels were nominally 18mg/mL in the strawberry (pH 8.29) and tobacco (pH 9.10) e-liquids and ranged between 3-18mg/mL in the usual brands (mean pH 6.80). Each day consisted of a 15-puff session followed by 4h of abstinence, then 90min of ad libitum use. Subjects used a KangerTech mini ProTank 3.ResultsAfter 15 puffs, the amount of nicotine inhaled and systemically retained were not significantly different between the strawberry and tobacco e-liquids but plasma AUC(0→180) was significantly higher with the strawberry e-liquid. While not significantly different, Cmax was 22% higher and various early time point AUCs to measure rate of rise of nicotine in blood ranged between 17 and 23% higher with the strawberry e-liquid compared to the tobacco e-liquid. During ad libitum use, systemic exposure to nicotine (AUC(0→90)) was the same for the tobacco and usual brand e-liquids but were both significantly lower than after using the strawberry e-liquid. The usual flavors were more liked and satisfying than the strawberry and tobacco e-liquids.ConclusionFlavors influence nicotine exposure through flavor liking, may affect rate of nicotine absorption possibly through pH effects, and contribute to heart rate acceleration and subjective effects of e-cigarettes. E-cigarette users titrate their nicotine exposure but the extent of titration may vary across flavors.

ObjectivesTo describe the effect of e-liquid flavors on nicotine intake and pharmacology of e-cigarettes.Methods11 males and 3 females participated in a 3-day inpatient crossover study with strawberry, tobacco, and their usual flavor e-liquid. Nicotine levels were nominally 18mg/mL in the strawberry (pH 8.29) and tobacco (pH 9.10) e-liquids and ranged between 3-18mg/mL in the usual brands (mean pH 6.80). Each day consisted of a 15-puff session followed by 4h of abstinence, then 90min of ad libitum use. Subjects used a KangerTech mini ProTank 3.ResultsAfter 15 puffs, the amount of nicotine inhaled and systemically retained were not significantly different between the strawberry and tobacco e-liquids but plasma AUC(0→180) was significantly higher with the strawberry e-liquid. While not significantly different, Cmax was 22% higher and various early time point AUCs to measure rate of rise of nicotine in blood ranged between 17 and 23% higher with the strawberry e-liquid compared to the tobacco e-liquid. During ad libitum use, systemic exposure to nicotine (AUC(0→90)) was the same for the tobacco and usual brand e-liquids but were both significantly lower than after using the strawberry e-liquid. The usual flavors were more liked and satisfying than the strawberry and tobacco e-liquids.ConclusionFlavors influence nicotine exposure through flavor liking, may affect rate of nicotine absorption possibly through pH effects, and contribute to heart rate acceleration and subjective effects of e-cigarettes. E-cigarette users titrate their nicotine exposure but the extent of titration may vary across flavors.

Thirdhand smoke (THS) is the fraction of cigarette smoke that persists in indoor environments after smoking. We investigated the effects of neonatal and adult THS exposure on bodyweight and blood cell populations in C57BL/6 J mice. At the end of neonatal exposure, THS-treated male and female mice had significantly lower bodyweight than their respective control mice. However, five weeks after neonatal exposure ended, THS-treated mice weighed the same as controls. In contrast, adult THS exposure did not change bodyweight of mice. On the other hand, both neonatal and adult THS exposure had profound effects on the hematopoietic system. Fourteen weeks after neonatal THS exposure ended, eosinophil number and platelet volume were significantly higher, while hematocrit, mean cell volume, and platelet counts were significantly lower compared to control. Similarly, adult THS exposure also decreased platelet counts and increased neutrophil counts. Moreover, both neonatal and adult THS exposure caused a significant increase in percentage of B-cells and significantly decreased percentage of myeloid cells. Our results demonstrate that neonatal THS exposure decreases bodyweight and that THS exposure induces persistent changes in the hematopoietic system independent of age at exposure. These results also suggest that THS exposure may have adverse effects on human health.

IntroductionElectronic cigarettes (e-cigarettes) are purported to deliver nicotine aerosol without any toxic combustion products present in tobacco smoke. In this longitudinal within-subjects observational study, we evaluated the effects of e-cigarettes on nicotine delivery and exposure to selected carcinogens and toxicants.MethodsWe measured seven nicotine metabolites and 17 tobacco smoke exposure biomarkers in the urine samples of 20 smokers collected before and after switching to pen-style M201 e-cigarettes for 2 weeks. Biomarkers were metabolites of 13 major carcinogens and toxicants in cigarette smoke: one tobacco-specific nitrosamine (NNK), eight volatile organic compounds (1,3-butadiene, crotonaldehyde, acrolein, benzene, acrylamide, acrylonitrile, ethylene oxide, and propylene oxide), and four polycyclic aromatic hydrocarbons (naphthalene, fluorene, phenanthrene, and pyrene). Changes in urine biomarkers concentration were tested using repeated measures analysis of variance.ResultsIn total, 45% of participants reported complete abstinence from cigarette smoking at 2 weeks, while 55% reported continued smoking. Levels of total nicotine and some polycyclic aromatic hydrocarbon metabolites did not change after switching from tobacco to e-cigarettes. All other biomarkers significantly decreased after 1 week of using e-cigarettes (p < .05). After 1 week, the greatest percentage reductions in biomarkers levels were observed for metabolites of 1,3-butadiene, benzene, and acrylonitrile. Total NNAL, a metabolite of NNK, declined by 57% and 64% after 1 and 2 weeks, respectively, while 3-hydroxyfluorene levels declined by 46% at week 1, and 34% at week 2.ConclusionsAfter switching from tobacco to e-cigarettes, nicotine exposure remains unchanged, while exposure to selected carcinogens and toxicants is substantially reduced.ImplicationsTo our knowledge, this is the first study that demonstrates that substituting tobacco cigarettes with an e-cigarette may reduce user exposure to numerous toxicants and carcinogens otherwise present in tobacco cigarettes. Data on reduced exposure to harmful constituents that are present in tobacco cigarettes and e-cigarettes can aid in evaluating e-cigarettes as a potential harm reduction device.

Thirdhand smoke (THS) is the contamination that persists after secondhand tobacco smoke has been emitted into air. It refers to the tobacco-related gases and particles that become embedded in materials, such as the carpet, walls, furniture, blankets, and toys. THS is not strictly smoke, but chemicals that adhere to surfaces from which they can be released back into the air, undergo chemical transformations and/or accumulate. Currently, the hazards of THS are not as well documented as the hazards of secondhand smoke (SHS). In this Perspective, we describe the distribution and chemical changes that occur as SHS is transformed into THS, studies of environmental contamination by THS, human exposure studies, toxicology studies using animal models and in vitro systems, possible approaches for avoiding exposure, remediation of THS contamination, and priorities for further research.

BackgroundTobacco has regained the status of the world's number two killer behind heart/vascular disease. Thirdhand smoke (THS) residue and particles from secondhand smoke (SHS) are suspected health hazards (eg, DNA damage) that are likely to contribute to morbidity and mortality, especially in vulnerable children. THS is easily transported and deposited indoors, where it persists and exposes individuals for months, creating potential health consequences in seemingly nicotine-free environments, particularly for vulnerable patients. We collected THS data to estimate infant exposure in the neonatal ICU (NICU) after visits from household smokers. Infant exposure to nicotine, potentially from THS, was assessed via assays of infant urine.MethodsParticipants were mothers who smoked and had an infant in the NICU (N=5). Participants provided surface nicotine samples from their fingers, infants' crib/incubator and hospital-provided furniture. Infant urine was analysed for cotinine, cotinine's major metabolite: trans-3'-hydroxycotinine (3HC) and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), a metabolite of the nicotine-derived and tobacco-specific carcinogen 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK).ResultsIncubators/cribs and other furniture had detectable surface nicotine. Detectable levels of cotinine, 3HC and NNAL were found in the infants' urine.DiscussionTHS appears to be ubiquitous, even in closely guarded healthcare settings. Future research will address potential health consequences and THS-reduction policies. Ultimately, hospital policies and interventions to reduce THS transport and exposure may prove necessary, especially for immunocompromised children.

ObjectiveTo characterize vaping behavior and nicotine intake during ad libitum e-cigarette access.MethodsThirteen adult e-cigarette users had 90 minutes of videotaped ad libitum access to their usual e-cigarette. Plasma nicotine was measured before and every 15 minutes after the first puff; subjective effects were measured before and after the session.ResultsAverage puff duration and interpuff interval were 3.5±1.4 seconds (±SD) and 118±141 seconds, respectively. 12% of puffs were unclustered puffs while 43%, 28%, and 17% were clustered in groups of 2-5, 6-10, and >10 puffs, respectively. On average, 4.0±3.3 mg of nicotine was inhaled; the maximum plasma nicotine concentration (Cmax) was 12.8±8.5 ng/mL. Among the 8 tank users, number of puffs was positively correlated with amount of nicotine inhaled, Cmax, and area under the plasma nicotine concentration-time curve (AUC0→90min) while interpuff interval was negatively correlated with Cmax and AUC0→90.ConclusionVaping patterns differ from cigarette smoking. Plasma nicotine levels were consistent with intermittent dosing of nicotine from e-cigarettes compared to the more bolus dosing from cigarettes. Differences in delivery patterns and peak levels of nicotine achieved could influence the addictiveness of e-cigarettes compared to conventional cigarettes.

ObjectiveThe rate of nicotine metabolism, determined primarily by CYP2A6 activity, influences tobacco dependence and smoking-induced disease risk. The prevalence of CYP2A6 gene variants differs by race, with greater numbers in African Americans compared with Caucasians. We studied nicotine disposition kinetics and metabolism by the CYP2A6 genotype and enzymatic activity, as measured by the nicotine metabolite ratio (NMR), in African American smokers.MethodsParticipants were administered intravenous infusions of deuterium-labeled nicotine and cotinine. Plasma and urine concentrations of nicotine and metabolites were measured and pharmacokinetic parameters were estimated.ResultsPharmacokinetic parameters and urine metabolite excretion data were analyzed by CYP2A6 genotype and by NMR. A number of gene variants were associated with markedly reduced nicotine and cotinine clearances. NMR was strongly correlated with nicotine (r=0.72) and cotinine (r=0.80) clearances. Participants with higher NMR excreted significantly greater nicotine C-oxidation and lower non-C-oxidation products compared with lower NMR participants.ConclusionCYP2A6 genotype, NMR, and nicotine pharmacokinetic data may inform studies of individual differences in smoking behavior and biomarkers of nicotine exposure.

BackgroundRate of nicotine metabolism is an important factor influencing cigarette smoking behavior, dependence, and efficacy of nicotine replacement therapy. The current study examined the hypothesis that chronic alcohol abuse can accelerate the rate of nicotine metabolism. Nicotine metabolite ratio (NMR, a biomarker for rate of nicotine metabolism) and patterns of nicotine metabolites were assessed at three time points after alcohol cessation.MethodsParticipants were 22 Caucasian men randomly selected from a sample of 165 smokers entering a 7-week alcohol dependence treatment program in Poland. Data were collected at three time points: baseline (week 1, after acute alcohol detoxification), week 4, and week 7. Urine was analyzed for nicotine and metabolites and used to determine the nicotine metabolite ratio (NMR, a biomarker for rate of nicotine metabolism), and total nicotine equivalents (TNE, a biomarker for total daily nicotine exposure).Results and conclusionsThere was a significant decrease in urine NMR over the 7 weeks after alcohol abstinence (F(2,42)=18.83, p

We tested the toxicity of thirdhand smoke (THS) using two controlled laboratory exposure scenarios and low levels of THS. One exposure modeled THS in a car parked outdoors, while the second modeled THS in a room without sunlight. The fabrics were exposed to cigarette smoke and then extracted in culture medium. Concentrations of nicotine, nicotine related alkaloids, and tobacco specific nitrosamines (TSNAs) were determined in fresh and aged extracts. The concentration of TSNAs increased with aging in the indoor experiment. THS extracts were used for cytotoxicity testing using mouse neural stem cells (mNSC), human dermal fibroblasts (hDF) and human palatal mesenchyme cells (hPM). Extracts from the car experiment inhibited mNSC proliferation in a live cell imaging assay and induced single strand DNA breaks in mNSC and hDF. In the indoor experiment, THS extracts made with medium containing serum proteins were significantly more toxic than extracts made with basal medium, and mNSC and hPM were more sensitive than hDF. These data indicate that: (1) aging of THS chemical differs on different fabrics and differs with and without sunlight; (2) very few cigarettes are sufficient to produce a toxic THS residue; and (3) protein enhances the efficiency of extraction of cytotoxic chemicals.

AimsTo measure the systemic retention of nicotine, propylene glycol (PG) and vegetable glycerin (VG) in electronic cigarette (e-cigarette) users, and assess the abuse liability of e-cigarettes by characterizing nicotine pharmacokinetics.DesignE-cigarette users recruited over the internet participated in a 1-day research ward study. Subjects took 15 puffs from their usual brand of e-cigarette. Exhaled breath was trapped in gas-washing bottles and blood was sampled before and several times after use.SettingSan Francisco, California, USA.ParticipantsThirteen healthy, experienced adult e-cigarette users (six females and seven males).MeasurementsPlasma nicotine was analyzed by gas chromatography-mass spectrometry (GC-MS/MS) and nicotine, VG and PG in e-liquids and gas traps were analyzed by LC-MS/MS. Heart rate changes and subjective effects were assessed.FindingsE-cigarettes delivered an average of 1.33 (0.87-1.79) mg [mean and 95% confidence interval (CI)] of nicotine, and 93.8% of the inhaled dose, 1.22 (0.80-1.66) was systemically retained. Average maximum plasma nicotine concentration (Cmax ) was 8.4 (5.4-11.5) ng/ml and time of maximal concentration (Tmax ) was 2-5 minutes. One participant had Tmax of 30 minutes. 84.4% and 91.7% of VG and PG, respectively, was systemically retained. Heart rate increased by an average of 8.0 beats per minute after 5 minutes. Withdrawal and urge to smoke decreased and the e-cigarettes were described as satisfying.ConclusionsE-cigarettes can deliver levels of nicotine that are comparable to or higher than typical tobacco cigarettes, with similar systemic retention. Although the average maximum plasma nicotine concentration in experienced e-cigarette users appears to be generally lower than what has been reported from tobacco cigarette use, the shape of the pharmacokinetic curve is similar, suggesting addictive potential.

BackgroundDifferences in carcinogen exposure from different cigarette products could contribute to differences in smoking-associated cancer incidence among Chinese compared with US smokers.MethodsUrine concentrations of metabolites of nicotine, the tobacco-specific nitrosamine (TSNA) 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), and polycyclic aromatic hydrocarbon metabolites (PAHs) were compared in 238 Chinese and 203 US daily smokers.ResultsComparing Chinese versus US smokers, daily nicotine intake and nicotine intake per cigarette smoked were found to be similar. When normalised for cigarettes per day, urine NNAL excretion was fourfold higher in US smokers, while the excretion of urine metabolites of the PAHs fluorene, phenanthrene and pyrene metabolites was 50% to fourfold higher in Chinese smokers (all, p

Background and aimsA broadly mandated reduction of the nicotine content (RNC) of cigarettes has been proposed in the United States to reduce the addictiveness of cigarettes, to prevent new smokers from becoming addicted and to facilitate quitting in established smokers. The primary aim of this study was to determine whether following 7 months of smoking very low nicotine content cigarettes (VLNC), and then returning to their own cigarettes, smokers would demonstrate persistently reduced nicotine intake compared with baseline or quit smoking.MethodsIn a community-based clinic 135 smokers not interested in quitting were randomized to one of two groups. A research group smoked their usual brand of cigarettes, followed by five types of research cigarettes with progressively lower nicotine content, each for 1 month, followed by 6 months at the lowest nicotine level (0.5 mg/cigarette) (53 subjects) and then 12 months with no intervention (30 subjects completed). A control group smoked their usual brand for the same period of time (50 subjects at 6 months, 38 completed). Smoking behavior, biomarkers of nicotine intake and smoke toxicant exposure were measured.ResultsAfter 7 months smoking VLNC, nicotine intake remained below baseline (plasma cotinine 149 versus 250 ng/ml, P

ObjectiveThe association between cigarette smoke exposure and the acute respiratory distress syndrome in patients with the most common acute respiratory distress syndrome risk factors of sepsis, pneumonia, and aspiration has not been well studied. The goal of this study was to test the association between biomarker-confirmed cigarette smoking and acute respiratory distress syndrome in a diverse cohort.DesignProspective cohort.SettingTertiary care center.PatientsFour hundred twenty-six critically ill patients with acute respiratory distress syndrome risk factors (excluding trauma and transfusion)Interventions: None.Measurements and main resultsWe obtained smoking histories and measured urine 4-(methylnitrosamino)-1-(3-pyridyl)-1- butanol (a biomarker of cigarette smoke exposure) on urine samples obtained at the time of study enrollment. The association between cigarette smoke exposure and acute respiratory distress syndrome differed based on acute respiratory distress syndrome risk factor (p < 0.02 for interaction). In patients with nonpulmonary sepsis as the primary acute respiratory distress syndrome risk factor (n = 212), 39% of those with acute respiratory distress syndrome were current smokers by history compared with 22% of those without acute respiratory distress syndrome (odds ratio, 2.28; 95% CI, 1.24-4.19; p = 0.008). Likewise, cigarette smoke exposure as measured by urine 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol was significantly associated with acute respiratory distress syndrome in this group. The increased risk of acute respiratory distress syndrome in nonpulmonary sepsis was restricted to patients with 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol levels consistent with active smoking and was robust to adjustment for other acute respiratory distress syndrome predictors. Cigarette smoke exposure as measured by history or 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol was not associated with acute respiratory distress syndrome in patients with other risk factors (e.g., pneumonia and aspiration).ConclusionsCigarette smoking measured both by history and biomarker is associated with an increased risk of acute respiratory distress syndrome in patients with nonpulmonary sepsis. This finding has important implications for tobacco product regulation and for understanding the pathogenesis of acute respiratory distress syndrome.