SIGNIFICANCE: Nicotine-containing products, labelled as being 'tobacco-free' nicotine (TFN), are marketed to consumers as alternatives to conventional tobacco products. Little is known about these emerging products and their contents. METHODS: Moisture, total nicotine and pH content were analysed in 70 commercially available TFN products, covering five different types (lozenges, chewing gum, loose leaf, toothpicks and pouches). The freebase nicotine was calculated using the measured pH values. RESULTS: Total nicotine levels ranged from 0.822 to 31.5 mg/g. Nicotine levels were highest in nicotine pouches (1.41-8.11 mg/product) and lowest in toothpicks (1.19-1.57 mg/product). Nicotine levels in TFN loose leaf (1.26-9.16 mg/g) were comparable to conventional moist snuff. The pH ranged from pH 4.68 to 9.49 and per cent freebase nicotine ranged from 0.0453% to 96.7%. The freebase nicotine content was highest in nicotine pouches (2.15-16.8 mg/g) and lowest in lozenges (0.0004-0.349 mg/g). The majority of TFN products (91.4%) analysed were advertised to contain flavour components. CONCLUSION: Overall, products advertised as higher strength were found to have higher nicotine content than products advertised as lower strength. The measured total nicotine content was either equal to or less than the level stated on the label, except for one product. Although TFN products may not contain tobacco lamina and may lack many harmful chemicals and carcinogens found in conventional smokeless products, freebase nicotine levels in the pouch products are elevated and could contribute to higher levels of addiction and other negative health effects.

The Centers for Disease Control and Prevention (CDC) Radiation Laboratory’s primary mission is to provide laboratory support for an effective and efficient response to public health radiological emergencies. The laboratory has developed methods for several radiological threat agents, including Iridium-192 (Ir-192). Ir-192 can be analyzed via its gamma energy through analytical methods such as High Purity Germanium (HPGe) and its beta energy through Liquid Scintillation Counting (LSC). In this work, we present and compare HPGe and LSC rapid response methods for Ir-192 quantification. Both methods show the reasonable results and can be used in emergency situations. © This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply 2024.

The kidneys, heart and lungs are vital organ systems evaluated as part of acute or chronic toxicity assessments. New methodologies are being developed to predict these adverse effects based on in vitro and in silico approaches. This paper reviews the current state of the art in predicting these organ toxicities. It outlines the biological basis, processes and endpoints for kidney toxicity, pulmonary toxicity, respiratory irritation and sensitization as well as functional and structural cardiac toxicities. The review also covers current experimental approaches, including off-target panels from secondary pharmacology batteries. Current in silico approaches for prediction of these effects and mechanisms are described as well as obstacles to the use of in silico methods. Ultimately, a commonly accepted protocol for performing such assessment would be a valuable resource to expand the use of such approaches across different regulatory and industrial applications. However, a number of factors impede their widespread deployment including a lack of a comprehensive mechanistic understanding, limited in vitro testing approaches and limited in vivo databases suitable for modeling, a limited understanding of how to incorporate absorption, distribution, metabolism, and excretion (ADME) considerations into the overall process, a lack of in silico models designed to predict a safe dose and an accepted framework for organizing the key characteristics of these organ toxicants.

Liquid Scintillation Counting (LSC) gross alpha/beta screening is a valuable tool for providing rapid laboratory response for the analysis of human clinical urine samples during a large-scale radiation incident event. Verification of method performance, as required for clinical laboratory testing, is accomplished by the evaluation of routine, periodic measurements of radioactive spiked samples for quality control, performance testing, and accuracy checks. Radionuclide stability of alpha and beta emitters in urine for LSC analysis is an important consideration. The purpose of this work is to demonstrate optimal preparations and storage conditions of samples used for method verification. © This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply 2024.

We measured levels of nitrosonornicotine (NNN) and 4-[methyl(nitroso)amino]-1-(3-pyridinyl)-1-butanone (NNK), the two most carcinogenic tobacco-specific nitrosamines, in the filler, binder, and wrapper of 50 cigars: 19 large cigars, 23 cigarillos, and 8 little cigars. The average NNN and NNK levels were 10.6 and 3.70 μg/g, respectively. These levels are 5- and 7-fold higher, respectively, than those of commercial cigarettes. The differences in NNN and NNK levels between cigars and cigarettes reflect differences in tobacco blends and tobacco treatments, such as fermentation. The average tobacco NNN and NNK levels of large cigars were 3- and 5-fold higher than those of cigarillos and little cigars, respectively. Large cigars also exhibited a significantly broader range of NNN and NNK than cigarillos and little cigars. The NNN and NNK levels in cigarillos are comparable to those of little cigars. These results are consistent with earlier studies finding that cigarillos and little cigars have similar tobacco blends with lower NNN and NNK content than large cigar tobacco blends.

This study described spatiotemporal changes in health insurance coverage, healthcare access, and reasons for non-insurance among racial/ethnic minority populations in the United States during the COVID-19 pandemic using four national survey datasets. Getis-Ord Gi* statistic and scan statistics were used to analyze geospatial clusters of health insurance coverage by race/ethnicity. Logistic regression was used to estimate odds of reporting inability to access healthcare across two pandemic time periods by race/ethnicity. Racial/ethnic differences in insurance were observed from 2010 through 2019, with the lowest rates being among Hispanic/Latino, African American, American Indian/Alaska Native, and Native Hawaiian/Pacific Islander populations. Pre-pandemic insurance coverage rates were geographically clustered. The percentage of adults citing change in employment status as the reason for non-insurance increased by about 7% after the start of the pandemic, with a small decrease observed among African American adults. Almost half of adults reported reduced healthcare access in June 2020, with 38.7% attributing reduced access to the pandemic; however, by May 2021, the percent of respondents reporting reduced access for any reason and due to the pandemic fell to 26.9% and 12.7%, respectively. In general, racial/ethnic disparities in health insurance coverage and healthcare access worsened during the pandemic. Although coverage and access improved over time, pre-COVID disparities persisted with African American and Hispanic/Latino populations being the most affected by insurance loss and reduced healthcare access. Cost, unemployment, and eligibility drove non-insurance before and during the pandemic.

Research gaps exist in toxic metals characterization in e-cigarette, or vaping, products (EVPs) as these analytes typically have low concentrations and most standard aerosol trapping techniques have high metals background. An additional complication arises from differences in the EVP liquid formulations with nicotine products having polar properties and non-nicotine products often being non-polar. Differences in polar and non-polar matrices and the subsequent aerosol chemistries from various EVPs required modifications of our previously reported nicotine-based EVP aerosol method. Validation and application of the expanded method, suitable for both hydrophobic and hydrophilic aerosols, are reported here. The metals analyzed for this study were Al, Cr, Fe, Co, Ni, Cu, Cd, Sn, Ba, and Pb. The method limits of detection for the modified method ranged from 0.120 ng/10 puffs for Cd to 29.3 ng/10 puffs for Al and were higher than reported for the previous method. Results of the analyses for metals in aerosols obtained from 50 EVP products are reported. Cannabinoid based EVP aerosols were below reportable levels, except for one sample with 16.08 ng/10 puffs for Cu. Nicotine-based EVP results ranged from 6.72 ng/10 puffs for Pb to 203 ng/10 puffs for Sn. Results of the analyses for these metals showed that aerosols from only 5 of the 50 devices tested had detectable metal concentrations. Concentrations of toxic elements in the aerosols for nicotine-based EVP aerosol metal concentration ranges were consistent with previously published results of aerosol analyses from this class of devices.

The present publication surveys several applications of in silico (i.e., computational) toxicology approaches across different industries and institutions. It highlights the need to develop standardized protocols when conducting toxicity-related predictions. This contribution articulates the information needed for protocols to support in silico predictions for major toxicological endpoints of concern (e.g., genetic toxicity, carcinogenicity, acute toxicity, reproductive toxicity, developmental toxicity) across several industries and regulatory bodies. Such novel in silico toxicology (IST) protocols, when fully developed and implemented, will ensure in silico toxicological assessments are performed and evaluated in a consistent, reproducible, and well-documented manner across industries and regulatory bodies to support wider uptake and acceptance of the approaches. The development of IST protocols is an initiative developed through a collaboration among an international consortium to reflect the state-of-the-art in in silico toxicology for hazard identification and characterization. A general outline for describing the development of such protocols is included and it is based on in silico predictions and/or available experimental data for a defined series of relevant toxicological effects or mechanisms. The publication presents a novel approach for determining the reliability of in silico predictions alongside experimental data. In addition, we discuss how to determine the level of confidence in the assessment based on the relevance and reliability of the information.

Hepatotoxicity is one of the most frequently observed adverse effects resulting from exposure to a xenobiotic. For example, in pharmaceutical research and development it is one of the major reasons for drug withdrawals, clinical failures, and discontinuation of drug candidates. The development of faster and cheaper methods to assess hepatotoxicity that are both more sustainable and more informative is critically needed. The biological mechanisms and processes underpinning hepatotoxicity are summarized and experimental approaches to support the prediction of hepatotoxicity are described, including toxicokinetic considerations. The paper describes the increasingly important role of in silico approaches and highlights challenges to the adoption of these methods including the lack of a commonly agreed upon protocol for performing such an assessment and the need for in silico solutions that take dose into consideration. A proposed framework for the integration of in silico and experimental information is provided along with a case study describing how computational methods have been used to successfully respond to a regulatory question concerning non-genotoxic impurities in chemically synthesized pharmaceuticals.

In May 2019, the Food and Drug Administration issued approval for Dengvaxia (Sanofi Pasteur), a live-attenuated, chimeric tetravalent dengue vaccine (1). In June 2021, the Advisory Committee on Immunization Practices (ACIP) recommended vaccination with Dengvaxia for children and adolescents aged 9–16 years with laboratory confirmation of previous dengue virus infection and who live in areas with endemic dengue transmission, such as the U.S. Virgin Islands (USVI)† (2). Confirming previous dengue virus infection before vaccine administration (prevaccination screening) is important because 1) although Dengvaxia decreases hospitalization and severe disease from dengue among persons with a previous infection, it increases the risk for these outcomes among persons without a previous infection; 2) many dengue virus infections are asymptomatic; and 3) many patients with symptomatic infections do not seek medical attention or receive appropriate testing (3). Sufficient laboratory evidence of previous dengue virus infection includes a history of laboratory-confirmed dengue§ or a positive serologic test result that meets ACIP-recommended performance standards for prevaccination screening, defined as high specificity (≥98%) and sensitivity (≥75%). A seroprevalence of 20% in the vaccine-eligible population (corresponding to a positive predictive value of ≥90% for a test with minimum sensitivity of 75% and minimum specificity of 98%) is recommended to maximize vaccine safety and minimize the risk for vaccinating persons without a previous dengue virus infection (2).

Little filtered cigars are tobacco products with many cigarette-like characteristics. However, despite cigars falling under the U.S. Food and Drug Administration regulatory authority, characterizing flavors, which are still allowed in little filtered cigars, and filter design may influence how people use the products and the resulting exposure to harmful and potentially harmful constituents. We estimated nicotine mouth level intake (MLI) from analyses of little cigar filter butt solanesol levels, brand characteristics, carbon monoxide boost, and puff volume in 48 dual cigarette/cigar users during two repeat bouts of ad lib smoking of three little filtered cigar brands. Mean nicotine MLI for the three brands was significantly different with Swisher Sweets (0.1% ventilation) Cherry at 1.20 mg nicotine, Cheyenne Menthol (1.5%) at 0.63 mg, and Santa Fe unflavored (49%) at 0.94 mg. The association between nicotine MLI and puff volume was the same between Cheyenne Menthol and Santa Fe unflavored. However, these were different from Swisher Sweets Cherry. At least five main factors─flavor, ventilation, filter design, nicotine delivery related to tar, and user puff volume─may directly or indirectly impact MLI and its association with other measures. We found that users of little filtered cigars that have different filter ventilation and flavor draw dissimilar amounts of nicotine from the product, which may be accompanied by differences in exposure to other harmful smoke constituents.

Acute toxicity in silico models are being used to support an increasing number of application areas including (1) product research and development, (2) product approval and registration as well as (3) the transport, storage and handling of chemicals. The adoption of such models is being hindered, in part, because of a lack of guidance describing how to perform and document an in silico analysis. To address this issue, a framework for an acute toxicity hazard assessment is proposed. This framework combines results from different sources including in silico methods and in vitro or in vivo experiments. In silico methods that can assist the prediction of in vivo outcomes (i.e., LD50) are analyzed concluding that predictions obtained using in silico approaches are now well-suited for reliably supporting assessment of LD50-based acute toxicity for the purpose of the Globally Harmonized System (GHS) classification. A general overview is provided of the endpoints from in vitro studies commonly evaluated for predicting acute toxicity (e.g., cytotoxicity/cytolethality as well as assays targeting specific mechanisms). The increased understanding of pathways and key triggering mechanisms underlying toxicity and the increased availability of in vitro data allow for a shift away from assessments solely based on endpoints such as LD50, to mechanism-based endpoints that can be accurately assessed in vitro or by using in silico prediction models. This paper also highlights the importance of an expert review of all available information using weight-of-evidence considerations and illustrates, using a series of diverse practical use cases, how in silico approaches support the assessment of acute toxicity. © 2022 Elsevier B.V.

Mainstream smoke yields of hydrogen cyanide (HCN) and three aromatic amines, 1-aminonaphthalene, 2-aminonaphthalene, and 4-aminobiphenyl, from 60 little cigar brands currently on the US market were measured for both International Organization for Standardization (ISO) and Canadian Intense (CI) smoking regimens. The smoke yields are compared with those from 50 cigarette products measured by Counts et al. of Philip Morris USA (PMUSA) in 2005 [Counts et al. Regul. Toxicol. Pharmacol. 2005 41, 185-227] and 50 cigarette products measured by the Centers for Disease Control and Prevention (CDC) in cooperation with the Food and Drug Administration (FDA) in 2012 [Tynan et al. Consumption of Cigarettes and Combustible Tobacco: United States, 2000-2011. In Morbidity and Mortality Weekly Report; Centers for Disease Control and Prevention, 2012; 565-580]. For the little cigars, the average HCN yield with the ISO smoking regimen is 335 μg/cigar (range: 77-809 μg/cigar), which is 332% higher than the average of 50 PMUSA 2005 cigarettes and 243% higher than the average of 50 CDC/FDA 2012 cigarettes. For the CI smoking regimen, the average HCN yield is 619 μg/cigar (range: 464-1045 μg/cigar), which is 70.5% higher than the average of 50 PMUSA 2005 cigarettes and 69% higher than the average of the 50 CDC/FDA 2012 cigarettes. For aromatic amines, the average ISO smoking regimen smoke yields are 36.6 ng/cigar (range: 15.9-70.6 ng/cigar) for 1-aminonaphthalene, 24.6 ng/cigar (range: 12.3-36.7 ng/cigar) for 2-aminonaphthalene, and 5.6 ng/cigar (range: 2.3-17.2 ng/cigar) for 4-aminobiphenyl. The average ISO yields of aromatic amines from little cigars are 141% to 210% higher compared to the average yields of 50 PMUSA cigarettes. The average CI smoke regimen yields are 73.0 ng/cigar (range: 32.1-112.2 ng/cigar) for 1-aminonaphthalene, 45.2 ng/cigar (range: 24.6-74.8 ng/cigar) for 2-aminonaphthalene, and 12.7 ng/cigar (range: 5.5-37.5 ng/cigar) for 4-aminobiphenyl. The average CI aromatic amine yields are 143% to 220% higher compared to the average yields of 50 PMUSA cigarettes, almost identical to the relative yields under the ISO smoking regimen. Both HCN and aromatic amine yields are 1.5× to 3× higher for the tested little cigars than for the conventional cigarettes; however, there are notable differences in the relationships of these yields to certain product characteristics, such as weight, ventilation, and tobacco type. The higher smoke yields of these compounds from little cigars indicates that cigar smokers may be at risk of a higher exposure to HCN and aromatic amines on a per stick basis and thus increased health concerns.

INTRODUCTION: In response to reducing cigarette nicotine content, people who smoke could attempt to compensate by using more cigarettes or by puffing on individual cigarettes with greater intensity. Such behaviors may be especially likely under conditions where normal nicotine content (NNC) cigarettes are not readily accessible. The current within-subject, residential study investigated whether puffing intensity increased with very low nicotine content (VLNC) cigarette use, relative to NNC cigarette use, when no other nicotine products were available. METHODS: Sixteen adults who smoke daily completed two 4-night hotel stays in Charleston, South Carolina (U.S.) in 2018 during which only NNC or only VLNC cigarettes were accessible. We collected the filters from all smoked cigarettes and measured the deposited solanesol to estimate mouth-level nicotine delivery per cigarette. These estimates were averaged within and across participants, per each 24-hour period. We then compared the ratio of participant-smoked VLNC and NNC cigarette mouth-level nicotine to the ratio yielded by cigarette smoking machines (when puffing intensity is constant). RESULTS: Average mouth-level nicotine estimates from cigarettes smoked during the hotel stays indicate participants puffed VLNC cigarettes with greater intensity than NNC cigarettes in each respective 24-hour period. However, this effect diminished over time (p<0.001). Specifically, VLNC puffing intensity was 40.0% (95% CI: 29.9, 53.0) greater than NNC puffing intensity in the first period, and 16.1% (95% CI: 6.9, 26.0) greater in the fourth period. CONCLUSION: Average puffing intensity per cigarette was elevated with exclusive VLNC cigarette use, but the extent of this effect declined across four days. IMPLICATIONS: In an environment where no other sources of nicotine are available, people who smoke daily may initially attempt to compensate for cigarette nicotine reduction by puffing on individual cigarettes with greater intensity. Ultimately, the compensatory behavior changes required to achieve usual nicotine intake from VLNC cigarettes are drastic and unrealistic. Accordingly, people are unlikely to sustain attempts to compensate for very low cigarette nicotine content.

BACKGROUND: Smokeless tobacco (ST) products are widely used throughout the world and contribute to morbidity and mortality in users through an increased risk of cancers and oral diseases. Bacterial populations in ST contribute to taste, but their presence can also create carcinogenic, Tobacco-Specific N-nitrosamines (TSNAs). Previous studies of microbial communities in tobacco products lacked chemistry data (e.g. nicotine, TSNAs) to characterize the products and identify associations between carcinogen levels and taxonomic groups. This study uses statistical analysis to identify potential associations between microbial and chemical constituents in moist snuff products. METHODS: We quantitatively analyzed 38 smokeless tobacco products for TSNAs using liquid chromatography with tandem mass spectrometry (LC-MS/MS), and nicotine using gas chromatography with mass spectrometry (GC-MS). Moisture content determinations (by weight loss on drying), and pH measurements were also performed. We used 16S rRNA gene sequencing to characterize the microbial composition, and additionally measured total 16S bacterial counts using a quantitative PCR assay. RESULTS: Our findings link chemical constituents to their associated bacterial populations. We found core taxonomic groups often varied between manufacturers. When manufacturer and flavor were controlled for as confounding variables, the genus Lactobacillus was found to be positively associated with TSNAs. while the genera Enteractinococcus and Brevibacterium were negatively associated. Three genera (Corynebacterium, Brachybacterium, and Xanthomonas) were found to be negatively associated with nicotine concentrations. Associations were also investigated separately for products from each manufacturer. Products from one manufacturer had a positive association between TSNAs and bacteria in the genus Marinilactibacillus. Additionally, we found that TSNA levels in many products were lower compared with previously published chemical surveys. Finally, we observed consistent results when either relative or absolute abundance data were analyzed, while results from analyses of log-ratio-transformed abundances were divergent.

Chemicals of unknown inhalational toxicity are present in electronic cigarette and vaping products. E-cigarettes typically contain nicotine and other relatively hydrophilic chemicals while vaping products typically contain cannabinoids and other hydrophobic chemicals. For example, vaping products can include hydrophobic terpenes such as squalane (SQA) and squalene (SQE). However, little is known about the SQA and SQE transmission from liquid to aerosol. SQA and SQE are used in commercial products that are applied dermally and ingested orally, but limited information is available on their inhalational exposure and toxicity. We developed and validated a quantitative method to measure SQE and SQA in bronchoalveolar lavage fluid to assess if these chemicals accumulate in lung epithelial lining fluid after inhalation. Calibration curves spanned a range of 0.50-30.0 µg analyte per mL bronchoalveolar lavage fluid. Recoveries were found to be 97-105% for SQE and 81-106% for SQA. Limits of detection were 0.50 μg/ml for both SQE and SQA. The method was applied to bronchoalveolar lavage fluid samples of patients from the 2019 outbreak of e-cigarette, or vaping, product use-associated lung injury (EVALI) and a comparison group. Neither SQA nor SQE was detected above the method LOD for any samples analyzed; conversely, SQA or SQE were reproducibly measured in spiked quality control BAL fluids (relative standards deviations <15% for both analytes). Further applications of this method may help to evaluate the potential toxicity of SQA and SQE chronically inhaled from EVPs.

A quantitative method was developed to measure four harmful carbonyls (acetaldehyde, acrolein, crotonaldehyde, and formaldehyde) in aerosol generated from e-cigarette, or vaping, products (EVPs). The method uses a commercially available sorbent bed treated with a derivatization solution to trap and stabilize reactive carbonyls in aerosol emissions from EVPs to reduce reactive analyte losses and improve quantification. Analytes were extracted from the sorbent material using acetonitrile and analyzed via high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). The method was applied to aerosols generated from products obtained from case patients with EVP use-associated lung injury (EVALI). The method accuracy ranged from 93.6 to 105% in the solvent and 99.0 to 112% in the matrix. Limits of detection (LODs) were in the low nanogram range at 0.735-2.10 ng for all analytes, except formaldehyde at 14.7 ng. Intermediate precision, as determined from the replicate measurements of quality-control (QC) samples, showed a relative standard deviation (RSD) of less than 20% for all analytes. The EVALI case-related products delivered aerosol containing the following ranges of carbonyls: acetaldehyde (0.0856-5.59 μg), acrolein (0.00646-1.05 μg), crotonaldehyde (0.00168-0.108 μg), and formaldehyde (0.0533-12.6 μg). At least one carbonyl analyte was detected in every product. Carbonyl deliveries from EVALI-associated products of all types are consistent with the previously published results for e-cigarettes, and levels are lower than those observed in smoke from combustible cigarettes. This method is rugged, has high throughput, and is well suited for quantifying four harmful carbonyls in aerosol emissions produced by a broad spectrum of devices/solvents, ranging from e-cigarette containing polar solvents to vaping products containing nonpolar solvents.

INTRODUCTION: Health-related behavioral risk factors and obesity are linked to high risk for multiple chronic diseases. We examined the prevalence of these risk factors among American Indians and Alaska Natives (AI/ANs) compared with that of non-Hispanic Whites and across Indian Health Service (IHS) regions. METHODS: We used 2017 Behavioral Risk Factor Surveillance System data from participants in 50 states and the District of Columbia to assess 4 behavioral risk factors (current cigarette smoking, heavy drinking, binge drinking, and physical inactivity) and obesity. We analyzed disparities in these risk factors between AI/AN and non-Hispanic White participants, nationwide and by IHS region, by conducting log-linear regression analyses while controlling for potential confounders. RESULTS: Nationwide, crude prevalence of current smoking, physical inactivity, and obesity were significantly higher among AI/AN than non-Hispanic White participants. After adjustment for sociodemographic characteristics, AI/AN participants were 11% more likely to report current smoking (P < .05) and 23% more likely to report obesity (P < .001) than non-Hispanic White participants. These patterns persisted in most IHS regions with some exceptions. In the Southwest region, AI/AN participants were 39% less likely to report current smoking than non-Hispanic White participants (P < .001). In the Pacific Coast region, compared with non-Hispanic White participants, AI/AN participants were 54% less likely to report heavy drinking (P < .01) but 34% more likely to report physical inactivity (P < .05). Across IHS regions, AI/AN participants residing in Alaska and the Northern Plains regions had the highest prevalence of current smoking and binge drinking, and those in the Southwest and Pacific Coast regions had the lowest prevalence of current smoking. AI/AN participants in the Southwest region had the lowest prevalence of physical inactivity, and those in the Southern Plains region had the highest prevalence of obesity. CONCLUSIONS: The findings of this study support the importance of public health efforts to address and improve behavioral risk factors related to chronic disease in AI/AN people, both nationwide and among IHS regions, through culturally appropriate interventions.

Numerous chemicals of unknown inhalational toxicity have been measured in electronic cigarette, or vaping, products (EVPs). In addition, little is known about the liquid-to-aerosol transmission and deliveries of these chemicals, including oil-like terpenes such as squalene (SQE) and squalane (SQA). To provide information on the aerosol deliveries of these compounds from EVPs, we developed and validated a quantitative method to measure squalene and squalane in EVP aerosol emissions. Validation parameters include measurement repeatability (SQA and SQE %RSD <6%), intermediate precision (SQA: %RSD 11%, SQE: %RSD 17%), accuracy (SQA: 86-107%, SQE: 104-113%), matrix effects, method robustness, and analyte stability. Limits of detection were 6.06 ng/mL puffed air volume for both squalene and squalane. The method was used to measure squalene and squalane in aerosol emissions of 153 EVPs associated with case patients from a recent outbreak of e-cigarette, or vaping, product use associated lung injury (EVALI). The EVPs analyzed were organized into nicotine, cannabidiol, and tetrahydrocannabinol products by the percentage of nicotine, cannabidiol, and tetrahydrocannabinol in total particulate matter after vaping. In case-associated tetrahydrocannabinol products the detection rates and mean concentrations were 82.4% and 33.0 ng/mL puffed air for squalene and 4.41% and 7.80 ng/mL puffed air for squalane.

We developed a quantitative method for analyzing nicotine and menthol in e-cigarette, or vaping, products (EVPs). These products may adversely impact health through inhalational exposure to addictive and harmful chemicals. The presence of unknown substances in do-it-yourself e-liquids, counterfeits, or unregulated products may increase exposure to harmful chemicals, as underscored by the 2019 EVP use-associated lung injury (EVALI) outbreak. To minimize these risks, it is important to accurately quantify nicotine and menthol in e-liquids and aerosol emissions to evaluate EVP authenticity, verify product label accuracy, and identify potentially hazardous products. We developed a simple, versatile, high-throughput method using isotope-dilution gas chromatography-mass spectrometry for quantifying nicotine and menthol concentrations in both e-liquid contents and machine-generated aerosol emissions of EVPs. Rigorous validation has demonstrated that the method is specific, precise (CV<2.71%), accurate (percent error ≤7.0%), and robust. Linear calibration ranges from 0.01 to 1.00 mg/ml for both analytes was achieved, corresponding to expected analyte levels in e-liquids and machine-generated EVP aerosols. Limits of detection (LODs) in the final 10-ml sample extract were 0.4 μg/ml for nicotine and 0.2 μg/ml for menthol. The method was used to analyze aerosol emissions of 141 EVPs associated with the 2019 EVALI outbreak; detectable levels of nicotine (2.19-59.5 mg/g of aerosol) and menthol (1.09-10.69 mg/g of aerosol) were observed in 28 and 11%, respectively, of the samples analyzed. Nicotine was not detected in any of the tetrahydrocannabinol (THC), cannabidiol (CBD), or oil-based products, while menthol (2.95 mg/g of aerosol) was only detected in one of these products (THC-labeled). The analytical method can be used to quantify nicotine and menthol concentrations in the e-liquids and aerosols from a range of EVPs, and these findings highlight a difference between e-cigarette and other vaping products.

Standard machine smoking protocols provide useful information for examining the impact of design parameters, such as filter ventilation, on mainstream smoke delivery. Unfortunately, their results do not accurately reflect human smoke exposure. Clinical research and topography devices in human studies yield insights into how products are used, but a clinical setting or smoking a cigarette attached to such a device may alter smoking behavior. To better understand smokers' use of filtered cigarette products in a more natural environment, we developed a low-cost, high-throughput approach to estimate mainstream cigarette smoke exposure on a per-cigarette basis. This approach uses an inexpensive flatbed scanner to scan smoked cigarette filter butts and custom software to analyze tar-staining patterns. Total luminosity, or optical staining density, of the scanned images provides quantitative information proportional to mainstream smoke-constituent deliveries on a cigarette-by-cigarette basis. Duplicate sample analysis using this new approach and our laboratory's gold-standard liquid chromatography/tandem mass spectrometry (LC/MS/MS) solanesol method yielded comparable results (+7% bias) from the analysis of 20 commercial cigarettes brands (menthol and nonmentholated). The brands varied in design parameters such as length, filter ventilation, and diameter. Plots correlating the luminosity to mainstream smoked-nicotine deliveries on a per-cigarette basis for these cigarette brands were linear (average R(2) > 0.91 for nicotine and R(2) > 0.83 for the tobacco-specific nitrosamine NNK), on a per-brand basis, with linearity ranging from 0.15 to 3.00 mg nicotine/cigarette. Analysis of spent cigarette filters allows exposures to be characterized on a per-cigarette basis or a "daily dose" via summing across results from all filter butts collected over a 24 h period. This scanner method has a 100-fold lower initial capital cost for equipment than the LC/MS/MS solanesol method and provides high-throughput results (~200 samples per day). Thus, this new method is useful for characterizing exposure related to filtered tobacco-product use.

The long-term health effects of using e-cigarette, or vaping, products (EVPs; also known as e-cigarettes, electronic nicotine delivery systems, and vape pens) remain largely unknown. The inhalation of excipients, such as propylene glycol (PG) and glycerin (GLY), may have long-term health effects. In addition to the direct health effects of PG and GLY, glycerin-containing products can be contaminated with toxic ethylene glycol (EG) and diethylene glycol (DEG). To assess this issue, we developed a simple, versatile, high-throughput isotope dilution gas chromatography-tandem mass spectrometry method for quantifying these common excipients and contaminants. The method is applicable to both the liquid contents and machine-generated aerosols of EVPs. Our rigorous method validation demonstrates that the new method is specific, precise, accurate, and rugged/robust. The calibration range is linear from 0.1-7 mg for the excipients and 2.5-1,000 µg for the contaminants. These ranges encompass expected excipients levels in EVP e-liquids and their machine-generated aerosols and the relevant maximum residue safety limit of 1 mg/g, or 0.1% (w/w), for the contaminants. The calculated limits of detection for PG, GLY, EG, and DEG were determined as 0.0109 mg, 0.0132 mg, 0.250 µg, and 0.100 µg, respectively. The method was applied to the aerosol emissions analysis of 141 EVPs associated with the 2019 lung injury outbreak, and found typical levels of PG (120.28-689.35 mg/g of aerosol) and GLY (116.83-845.96 mg/g of aerosol) in all nicotine-containing products; PG (81.58-491.92 mg/g of aerosol) and GLY (303.86-823.47 mg/g of aerosol) in 13% of cannabidiol (CBD) products; PG (74.02-220.18 mg/g of aerosol) and GLY (596.43-859.81 mg/g of aerosol) in products with neither nicotine nor CBD; and none detected in tetrahydrocannabinol (THC) products. No products contained glycol contaminants above the recommended maximum residue safety limit.

E-cigarette, or vaping, product (EVP) use has increased dramatically in the United States over the last 4 years, particularly in youth and young adults. Little information is available on the chemical contents of these products. Typically, EVPs contain an active ingredient such as nicotine, CBD, or THC dissolved in a suitable solvent that facilitates aerosol generation. One EVP solvent, vitamin E acetate (VEA), has been measured in EVP liquids associated with lung injury. However, no validated analytical methods for measuring VEA in the aerosol from these devices was previously available. Therefore, we developed a high throughput isotope dilution LC-MS/MS method to simultaneously measure VEA and three other related tocopherols in aerosolized EVP samples. The assay was precise, with VEA repeatability ranging from 4.0 to 8.3% and intermediate precision ranging from 2.5 to 6.7%. Similar precision was obtained for the three other tocopherols measured. The LODs for the four analytes ranged from 8.85 × 10(-6) to 2.28 × 10(-5) μg analyte per mL of aerosol puff volume, and calibration curves were linear (R (2) > 0.99). This method was used to analyze aerosol emissions of 147 EVPs associated with EVALI case patients. We detected VEA in 46% of the case-associated EVPs with a range of 1.87 × 10(-4)-74.1 µg per mL of aerosol puff volume and mean of 25.1 µg per mL of aerosol puff volume. Macro-levels of VEA (>0.1% w/w total aerosol particulate matter) were not detected in nicotine or cannabidiol (CBD) products; conversely 71% of the EVALI associated tetrahydrocannabinol (THC) products contained macro-levels of VEA. Trace levels of other tocopherol isoforms were detected at lower rates and concentrations (α-tocopherol: 41% detected, mean 0.095 µg analyte per mL of aerosol puff volume; γ-tocopherol: 5% detected, mean 0.0193 µg analyte per mL of aerosol puff volume; δ-tocopherol: not detected). Our results indicate that VEA can be efficiently transferred to aerosol by EVALI-associated EVPs vaped using a standardized protocol.

The most commonly observed forms of aluminum, silicon and titanium in tobacco products are aluminum silicates (e.g., kaolin), silica and titanium(IV) oxide. These compounds are neither water soluble nor volatile at cigarette combustion temperatures. Rather, they are transported in mainstream tobacco smoke as particles after being freed by combustion from the tobacco filler and can induce pulmonary inflammation when inhaled. Aluminum silicate particles are the most frequently observed particles in the pulmonary macrophages of smokers and have become known as 'smokers' inclusions'. A relatively new technique, single particle triple quadrupole inductively coupled plasma-mass spectrometry was used to analyze aluminum-, silicon- and titanium-containing particle deliveries in cigarette and little cigar mainstream tobacco smoke, and to collect information on solid inorganic particles. The mass concentration of aluminum-containing particles transmitted in mainstream smoke was low (0.89-0.56 ng/cigarette), which was not surprising because aluminum silicates are not volatile. Although the collective masses (ng/cigarette) of aluminum-, silicon- and titanium-containing particles under 100 nm diameter transported in mainstream smoke were low, an abundance of 'ultrafine' particles (particles < 100 nm or nanoparticles) was observed. Limitations of the particle background equivalent diameter (the smallest detectable particle size (MassHunter 4.5 Software) due to the environmentally ubiquitous silicon background restricted the determination of silica nanoparticles, but silica particles slightly below 200 nm diameter were consistently detected. Aluminum- and titanium-containing nanoparticles were observed in all cigarette and little cigar samples, with titanium(IV) oxide particle deliveries consistently fewer in number and smaller in diameter than the other two types of particles. The highest concentrations of aluminum-containing particles (as kaolin) were in the nanoparticle range with much lower concentrations extending to the larger particle sizes (>100 nm). The number and range of particle sizes determined in mainstream smoke is consistent with pulmonary deposition of aluminum silicates described by other researchers as contributing to the 'smokers' inclusions' observed in pulmonary macrophages.

Two-tail t test statistical analyses of International Organization for Standardization nonintense and Canadian Intense mainstream smoke yields of total particulate matter, tar, nicotine, and carbon monoxide from cigarettes show that mean quantities are generally higher for a linear smoking machine at a 95% confidence level but a rotary smoking machine has better precision. A novel "super pad" analysis concept combines four smaller filter pads from a linear smoking machine, resulting in increased mean constituent yields and reduced variability. Although measurement variability is still greater than that of rotary machines, super padding may be useful to reduce the variance caused by linear smoking machines.

INTRODUCTION: Nicotine pouch products, oral smokeless products that contain nicotine but no tobacco leaf material, have recently entered the US marketplace. Available data indicate sales of these products in the United States have increased since 2018; however, the extent of use among US youth and adults is uncertain. METHODS: To assay the chemistry of these emerging tobacco products, we analyzed 37 nicotine pouch brands from six total manufacturers. Almost all of the products had flavor descriptors (36 of 37), such as mint, licorice, coffee, cinnamon, and fruit. The amount of free nicotine, the form most easily absorbed, was calculated for each product using total nicotine, product pH, the appropriate pKa, and the Henderson-Hasselbalch equation. RESULTS: Nicotine pouch products varied in pouch content mass, moisture content (1.12%‒47.2%), alkalinity (pH 6.86‒10.1), and % free nicotine (7.7%‒99.2%). Total nicotine content ranged from 1.29 to 6.11 mg/pouch, whereas free nicotine ranged from 0.166 to 6.07 mg/pouch. These findings indicate that nicotine and pH levels found in some of these nicotine pouches are similar to conventional tobacco products, such as moist snuff and snus, and that most of these pouch products are flavored. CONCLUSIONS: Although these products likely lack many tobacco-related chemicals, each product analyzed contained nicotine, which is both addictive and can harm human health. Given that nicotine pouches may appeal to a spectrum of users, from novice to experienced users, it is important to include these emerging tobacco products in tobacco control research, policy, and practice. IMPLICATIONS: These "tobacco-free" nicotine pouches have similar pH and nicotine content to conventional tobacco products, such as moist snuff and snus. Although they lack many tobacco-related chemicals, most are highly flavored which could increase experimentation from new users. Given that nicotine pouches may appeal to a spectrum of users, from novice to experienced users, in terms of their flavors and nicotine content, it is important to examine and include these emerging tobacco products as they relate to tobacco control research, policy, and practice.

Solanesol, a naturally occurring constituent of tobacco, has been utilized as a good marker for environmental tobacco smoke particulate and as a non-invasive predictor of mainstream cigarette smoke tar and nicotine intake under naturalistic smoking conditions. A fast and accurate method for measuring free solanesol to assess tobacco smoke exposure is highly desirable. We have developed and validated a new environmentally friendly, high throughput method for measuring solanesol content in discarded cigarette filter butts. The solanesol deposited in the used filters can be correlated with mainstream smoke deliveries of nicotine and total particle matter (TPM) to estimate constituent delivery to smokers. A portion of filter material is removed from cigarette butts after machine smoking, spiked with internal standard solution, extracted, and quantitatively analyzed using reverse phase liquid chromatography coupled to a triple quadrupole mass spectrometer. The new method incorporates a 48-well plate format for automated sample preparation that reduces sample preparation time and solvent use and increases sample throughput 10-fold compared to our previous method. Accuracy and precision were evaluated by spiking known amounts of solanesol on both clean and smoked cigarette butts. Recoveries exceeded 93% at both low and high spiking levels. Linear solanesol calibration curves ranged from 1.9 to 367 µg/butt with a 0.05 µg/butt limit of detection.

We investigated the suitability of isotope-dilution liquid chromatography coupled with tandem mass spectrometry for identifying vitamin E acetate (VEA) in bronchoalveolar lavage (BAL) fluid. This new method demonstrates high accuracy, selectivity, and sensitivity, with mean recoveries higher than 90%, coefficients of variation ranging from 1.5% to 4.5%, and a limit of detection of 1.10 ng/mL. Calibration curves were linear (R(2) > 0.99). The linear range and detection limit of the method were adequate for identifying VEA in 48 of 51 BAL fluid samples collected from people with lung injury resulting from e-cigarettes, or vaping, product use. We conclude that this method is an effective tool for studying VEA accumulation in lungs caused by using e-cigarettes, or vaping, products that contain VEA.

Nitromethane is a volatile organic compound categorized as a Group 2B carcinogen by the International Agency for Research on Cancer. It has been detected in mainstream cigarette smoke, but few reliable methods have been reported for accurate quantification. We developed, a sensitive, selective, fully validated method for the targeted determination of nitromethane in mainstream tobacco smoke in ten U.S. domestic brands and two quality control materials (3R4F and CM6). The vapor phase portion of machine-generated cigarette mainstream smoke, under modified ISO 3308:2000 regime (ISO) and modified intense regime (HCI), from single cigarettes was collected using airtight polyvinylfluoride sampling bags. The bags' contents were extracted using methanol containing an isotopically labeled internal standard followed by gas chromatography-tandem mass spectrometry. This approach is sufficiently sensitive to measure nitromethane levels in the nanogram range, with a method limit of detection of 72.3 ng/cig. Within-product variability estimated from the replicate analysis of 10 products ranged from 4.6%-16.3% (n = 6) over the two different smoking regimes, and method reproducibility estimated from two products used as quality control materials (3R4F and CM6) yielded intermediate precision values ranging from 16.6 to 20.8% (n = 20). Under HCI, nitromethane yields in machine-generated cigarette smoke from ten different domestic cigarette products ranged from 3.2 to 12 μg/cig; under ISO yields ranged from 1.6 to 4.9 μg/cig under standardized smoking machine conditions. Nitromethane yields are related to both the smoke regime (blocking of vent holes, puff duration and puff volume) and the heterogeneity of tobacco mixtures. This method provides a selective and fully validated technique to accurately quantify nitromethane in mainstream cigarette smoke, with minimal waste generation. It is an improvement over previous methods with regards to specificity, throughput, and simplicity of the sample collection process.

PURPOSE: To examine the chemical composition of JUUL pods collected from a convenience sample of 16 high schools in California to identify possible consumer modification or counterfeit use. METHODS: Using Gas Chromatography-Mass Spectrometry, we quantitatively analyzed the nicotine, propylene glycol (PG), and vegetable glycerin (VG) in JUUL pods (n = 26) collected from California high schools and compared results to commercial 3% (n = 15) and 5% (n = 24) JUUL pods purchased online. RESULTS: Most of the collected JUUL pods (24/26 pods) had a nicotine concentration (43.3 mg/ml, 95% PI: 21.5-65.1) outside the prediction intervals (PI) of the 3% (33.5 mg/ml, 95% PI: 31.8-35.2) and 5% (55.0 mg/ml, 95% PI: 51.5-58.3) commercial JUUL pods. Most (73%) collected JUUL pods had VG concentrations (583.5 mg/ml, PI: 428.9-738.1) lower than the 3% (722.2 mg/ml, PI: 643.0-801.4) and 5% (710.5 mg/ml, PI: 653.1-767.8) commercial JUUL pods. CONCLUSIONS: Used JUUL products collected from high school students or found on school grounds were not chemically consistent with the manufacturer's stated formulations.