Electrically Heated Cigarette Smoking System Research Articles

Reduced exposure evaluation of an Electrically Heated Cigarette Smoking System. Part 8: Nicotine bridging – Estimating smoke constituent exposure by their relationships to both nicotine levels in mainstream cigarette smoke and in smokers

A modeling approach termed ‘nicotine bridging’ is presented to estimate exposure to mainstream smoke constituents. The method is based on: (1) determination of harmful and potentially harmful constituents (HPHC) and in vitro toxicity parameter-to-nicotine regressions obtained using multiple machine-smoking protocols, (2) nicotine uptake distributions determined from 24-h excretion of nicotine metabolites in a clinical study, and (3) modeled HPHC uptake distributions using steps 1 and 2. An example of ‘nicotine bridging’ is provided, using a subset of the data reported in Part 2 of this supplement (Zenzen et al., 2012) for two conventional lit-end cigarettes (CC) and the Electrically Heated Cigarette Smoking System (EHCSS) series-K6 cigarette. The bridging method provides justified extrapolations of HPHC exposure distributions that cannot be obtained for smoke constituents due to the lack of specific biomarkers of exposure to cigarette smoke constituents in clinical evaluations. Using this modeling approach, exposure reduction is evident when the HPHC exposure distribution curves between the MRTP and the CC users are substantially separated with little or no overlap between the distribution curves.

Reduced exposure evaluation of an Electrically Heated Cigarette Smoking System. Part 2: Smoke chemistry and in vitro toxicological evaluation using smoking regimens reflecting human puffing behavior

Chemical analysis of up to 49 harmful and potentially harmful constituents (HPHC) in mainstream smoke, in vitro cytotoxicity of the particulate and gas/vapor phase of mainstream smoke determined in the Neutral Red Uptake assay, and in vitro bacterial mutagenicity of the particulate phase determined in the Salmonella typhimurium Reverse Mutation (Ames) assay are reported for three Electrically Heated Cigarette Smoking System (EHCSS) series-K cigarettes, the University of Kentucky Reference Cigarette 2R4F, and a number of comparator commercial conventional lit-end cigarettes (CC) under ISO machine-smoking conditions and a total of 25 additional smoking regimens reflecting ‘human puffing behavior’ (HPB). The smoking machines were set to deliver nicotine yields for the EHCSS and comparator CC derived from the 10th percentile to the 90th percentile of nicotine uptake distributions in smokers determined in two clinical studies. Duplication of the smoking intensity ‘per cigarette’ on a smoking machine may provide an insight into product performance that is directly relevant to obtaining scientific evidence for reduced exposure substantiation based on mainstream cigarette smoke HPHC-to-nicotine regressions. The reported data support an overall evaluation of reduced exposure to HPHC and biological activity.

Reduced exposure evaluation of an Electrically Heated Cigarette Smoking System. Part 4: Eight-day randomized clinical trial in Korea

A randomized, controlled, open-label parallel-group, single-center study to determine biomarkers of exposure to 12 selected harmful and potentially harmful constituents (HPHC) in cigarette smoke and urinary excretion of mutagenic material in 72 male and female Korean subjects smoking Lark One cigarettes (1.0mg tar, 0.1mg nicotine, and 1.5mg CO) at baseline. Subjects were randomized to continue smoking Lark One cigarettes, or switch to an Electrically Heated Cigarette Smoking System (EHCSS) and EHCSS-K3 cigarette (3mg tar, 0.2mg nicotine, and 0.6mg CO), or to no-smoking. The mean decreases from baseline to Day 8 were statistically significant (all p<0.05) for 10 of 12 HPHC in mainstream cigarette smoke including CO (the primary objective) in the EHCSS-K3 group (range: −1.5% to −74.2%). Exposure to the other determined HPHC was not significantly different. In the Lark One group, the mean exposure to 6 of 12 HPHC in cigarette smoke was significantly (all p<0.05) decreased; however, exposure to CO was significantly increased. The largest mean reductions in biomarkers of exposure to HPHC occurred in smokers who switched to no-smoking (−3.4% to −98.9%). The mean excretion of mutagenic material was significantly decreased (p<0.05) in the EHCSS-K3 and no-smoking groups (−31.8% and −45.3%, respectively), and increased in the Lark One group (+31.5%).

Reduced exposure evaluation of an Electrically Heated Cigarette Smoking System. Part 3: Eight-day randomized clinical trial in the UK

A randomized, controlled, open-label, parallel-group, single-center study to determine biomarkers of exposure to nine selected harmful and potentially harmful constituents (HPHC) in cigarette smoke and urinary excretion of mutagenic material in 160 male and female subjects smoking Marlboro cigarettes (6mg tar, 0.5mg nicotine, and 7.0mg CO) at baseline. Subjects were randomized to continue smoking Marlboro cigarettes, or switch to using an Electrically Heated Cigarette Smoking System (EHCSS) smoking one of two EHCSS series-K cigarettes, the EHCSS-K6 cigarette (5mg tar, 0.3mg nicotine, and 0.6mg CO) or the EHCSS-K3 cigarette (3mg tar, 0.2mg nicotine, and 0.6mg CO), or switch to smoking Philip Morris One cigarettes (1mg tar, 0.1mg nicotine, and 2.0mg CO), or to no-smoking. The mean decreases from baseline to Day 8 were statistically significant (p⩽0.05) for all determined HPHC including benzene and CO (the primary objectives), and urinary excretion of mutagenic material in the EHCSS-K6 (range −35.5±29.2% to −79.4±14.6% [mean±standard deviation]), EHCSS-K3 (range −41.2±26.6% to −83.1±9.2%), and PM1 (range −14.6±24.1% to −39.4±17.5%) groups. The largest reductions in exposure occurred in the no-smoking group (range −55.4±45.0% to −100.0±0.0%).

Reduced exposure evaluation of an Electrically Heated Cigarette Smoking System. Part 5: 8-Day randomized clinical trial in Japan

A randomized, controlled, open-label, parallel-group, single-center study to determine biomarkers of exposure to twelve selected harmful and potentially harmful constituents (HPHCs) in cigarette smoke and urinary excretion of mutagenic material in 128 male and female Japanese subjects smoking Marlboro cigarettes (6mg tar, 0.5mg nicotine, and 7.0mg CO) at baseline. Subjects were randomized to continue smoking Marlboro cigarettes, or switch to the Electrically Heated Cigarette Smoking System (EHCSS) and smoke either the EHCSS-K6 (5mg tar, 0.3mg nicotine, and 0.6mg CO) or the EHCSS-K3 (3mg tar, 0.2mg nicotine, and 0.6mg CO) cigarette, or switch to smoking Lark One cigarettes (1mg tar, 0.1mg nicotine, and 2.0mg CO), or to no-smoking. The mean decreases from baseline to Day 8 were statistically significant (p⩽0.05) for all cigarette smoke HPHC including CO (the primary objective) and excretion of mutagenic material in the EHCSS-K6 (range: −14.6% to −75.6%) and EHCSS-K3 (range: −9.8% to −73.0%) groups. Statistically significant reductions (all p⩽0.05) in exposure to ten cigarette smoke HPHC (range: −5.9% to −34.6%), but not urinary mutagenicity, were observed in the Lark One group. The largest mean reductions in exposure to HPHC (all p⩽0.01 level) occurred in the no-smoking group (range: −13.7% to −97.6%).

Short term effects of reduced exposure to cigarette smoke on white blood cells, platelets and red blood cells in adult cigarette smokers

Previous studies indicate that cigarette smokers have a 5–30% higher white blood cell counts (WBC) compared to non-smokers and higher red blood cell counts. Methods: This study was to pool hematology data from three similar studies and analyze the data for effects on WBC, its subpopulations, platelets, red blood cell count (RBC) and hematocrit in adult cigarette smokers three days after using an electrically heated cigarette smoking system (EHCSS) as a potential reduced exposure product (PREP) or no-smoking compared to smoking a conventional cigarette. Results: Lower exposure to cigarette smoke in adult, long term smokers, by using an EHCSS or stopping smoking, leads to statistically significant decreases of up to 9% in WBC, neutrophils, lymphocytes, platelets, RBC and hematocrit within three days. Switching from CC-smoking to EHCSS-smoking or no-smoking resulted in lower WBC and vice versa within 3days. Conclusion: This clinical model may be used as a screening tool to find new technologies that could provide insights on changes in inflammation resulting from the change in cigarette smoke.

Determination of methyl-, 2-hydroxyethyl- and 2-cyanoethylmercapturic acids as biomarkers of exposure to alkylating agents in cigarette smoke

Alkylating agents occur in the environment and are formed endogenously. Tobacco smoke contains a variety of alkylating agents or precursors including, among others, N-nitrosodimethylamine (NDMA), 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), acrylonitrile and ethylene oxide. We developed and validated a method for the simultaneous determination of methylmercapturic acid (MMA, biomarker for methylating agents such as NDMA and NNK), 2-hydroxyethylmercapturic acid (HEMA, biomarker for ethylene oxide) and 2-cyanoethylmercapturic acid (CEMA, biomarker for acrylonitrile) in human urine using deuterated internal standards of each compound. The method involves liquid/liquid extraction of the urine sample, solid phase extraction on anion exchange cartridges, derivatization with pentafluorobenzyl bromide (PFBBr), liquid/liquid extraction of the reaction mixture and LC–MS/MS analysis with positive electrospray ionization. The method was linear in the ranges of 5.00–600, 1.00–50.0 and 1.50–900 ng/ml for MMA, HEMA and CEMA, respectively. The method was applied to two clinical studies in adult smokers of conventional cigarettes who either continued smoking conventional cigarettes, were switched to test cigarettes consisting of either an electrically heated cigarette smoking system (EHCSS) or having a highly activated carbon granule filter that were shown to have reduced exposure to specific smoke constituents, or stopped smoking. Urinary excretion of MMA was found to be unaffected by switching to the test cigarettes or stop smoking. Urinary HEMA excretion decreased by 46 to 54% after switching to test cigarettes and by approximately 74% when stopping smoking. Urinary CEMA excretion decreased by 74–77% when switching to test cigarettes and by approximately 90% when stopping smoking. This validated method for urinary alkylmercapturic acids is suitable to distinguish differences in exposure not only between smokers and nonsmokers but also between smoking of conventional and the two test cigarettes investigated in this study.

Comparison of Environmental Tobacco Smoke (ETS) Concentrations Generated by an Electrically Heated Cigarette Smoking System and a Conventional Cigarette

Smoking conventional lit-end cigarettes results in exposure of nonsmokers to potentially harmful cigarette smoke constituents present in environmental tobacco smoke (ETS) generated by sidestream smoke emissions and exhaled mainstream smoke. ETS constituent concentrations generated by a conventional lit-end cigarette and a newly developed electrically heated cigarette smoking system (EHCSS) that produces only mainstream smoke and no sidestream smoke emissions were investigated in simulated “office” and “hospitality” environments with different levels of baseline indoor air quality. Smoking the EHCSS (International Organisation for Standardization yields: 5 mg tar, 0.3 mg nicotine, and 0.6 mg carbon monoxide) in simulated indoor environments resulted in significant reductions in ETS constituent concentrations compared to when smoking a representative lit-end cigarette (Marlboro: 6 mg tar, 0.5 mg nicotine, and 7 mg carbon monoxide). In direct comparisons, 24 of 29 measured smoke constituents (83%) showed mean reductions of greater than 90%, and 5 smoke constituents (17%) showed mean reductions between 80% and 90%. Gas–vapor phase ETS markers (nicotine and 3-ethenylpyridine) were reduced by an average of 97% (range 94–99%). Total respirable suspended particles, determined by online particle measurements and as gravimetric respirable suspended particles, were reduced by 90% (range 82–100%). The mean and standard deviation of the reduction of all constituents was 94 ± 4%, indicating that smoking the new EHCSS in simulated “office” and “hospitality” indoor environments resulted in substantial reductions of ETS constituents in indoor air.

Environmental tobacco smoke (ETS) evaluation of a third-generation electrically heated cigarette smoking system (EHCSS)

This sub-study of a randomized, controlled, forced-switching, open-label, parallel-group, clinical study compared environmental tobacco smoke (ETS) produced when 60 male and female adult smokers switched to a third-generation electrically heated cigarette smoking system (EHCSS), continued to smoke a conventional cigarette (CC), or stopped smoking (No-smoking). Concentrations of air constituents including respirable suspended particulate (RSP), carbon monoxide (CO), ammonia and total volatile organic compounds (TVOCs) and ETS markers including solanesol-related particulate matter (Sol-PM), ultraviolet absorbing particulate matter (UVPM), fluorescent particulate matter (FPM), nicotine and 3-ethenyl pyridine (3-EP) were measured in a ventilated, furnished conference room over a 2-h period on separate occasions for each smoking condition. When the EHCSS was used, concentrations of CO and most ETS markers were in the same range as during no-smoking. Concentrations of ammonia were reduced by 41% and concentrations of other selected constituents of ETS were reduced by 87–99% in the air of a room in which EHCSS cigarettes were smoked as compared to concentrations in the same room when conventional cigarettes were smoked. Switching from conventional cigarette smoking to the EHCSS resulted in substantial reductions in concentrations of several markers of environmental tobacco smoke.

Short-term clinical exposure evaluation of a third-generation electrically heated cigarette smoking system (EHCSS) in adult smokers

This randomized, controlled, forced-switching, open-label, parallel-group, single-center study in 100 male and female adult smokers evaluated 12 biomarkers of tobacco smoke exposure. We measured exposure to the following smoke constituents: nicotine, pyrene, tobacco-specific nitrosamines, three aromatic amines, carbon monoxide, benzene, acrolein, crotonaldehyde, and 1,3-butadiene. After baseline exposure determination, adult smokers of a conventional cigarette (CC) were switched to an electrically heated cigarette smoking system (EHCSS, Series K), continued smoking the CC, or stopped smoking (No-smoking) for 8 days in a controlled, confined, clinical setting. In the EHCSS group, the mean decrease from Baseline to Day 8 in the biomarkers of exposure ranged from 16% to 77% at Day 8 compared to Baseline. After adjusting for the residual effect (carryover effects due to long elimination half-life and non-tobacco confounding sources of exposure), the mean percent decrease from Baseline for all 12 biomarkers ranged from 47% to 90%. In conclusion, switching for 8 days from a conventional cigarette to the EHCSS substantially reduced exposure of adult smokers to several constituents of both the particulate and gas phases of cigarette smoke.

12-Week clinical exposure evaluation of a third-generation electrically heated cigarette smoking system (EHCSS) in adult smokers

This randomized, controlled, forced-switching, open-label, parallel-group, single-center study in 90 male and female adult smokers evaluated six biomarkers of tobacco smoke exposure over a 12-week period of unrestricted smoking in the participants’ normal life setting. Baseline biomarker levels were measured, then participants were randomly assigned to switch to an electrically heated cigarette smoking system (EHCSS, Series K) or to continue smoking a conventional cigarette (CC) of similar tar yield (Federal Trade Commission method) for 12 weeks. Compared to Baseline, adult smokers who switched to the EHCSS for 12 weeks in their normal life setting had significantly reduced nicotine equivalents (−33%), total NNAL (a biomarker for NNK, −63%), 1-OHP (a surrogate biomarker for polycyclic aromatic hydrocarbons, −38%), carboxyhemoglobin (a biomarker for carbon monoxide, −23%), 3-HPMA (a biomarker for acrolein, −25%) and S-PMA (a biomarker for benzene, −49%), whereas exposure was stable in the CC control group.

A 12‐Month, Randomized, Controlled Study to Evaluate Exposure and Cardiovascular Risk Factors in Adult Smokers Switching From Conventional Cigarettes to a Second‐Generation Electrically Heated Cigarette Smoking System

This randomized, controlled, forced-switching, open-label, parallel-group study in 97 adult male and female smokers of conventional cigarettes evaluated biomarkers of tobacco smoke exposure and cardiovascular risk factors. After baseline measurements, smokers were either switched to a second-generation electrically heated cigarette smoking system (EHCSS) or continued smoking conventional cigarettes for 12 months. Biomarkers of exposure and cardiovascular risk factors were measured at 0.5, 1, 2, 3, 4, 5, 6, 9, and 12 months. There was a rapid and sustained reduction in all biomarkers of exposure after switching to the EHCSS, with statistically significant reductions from baseline in nicotine equivalents (-18%), plasma cotinine (-16%), total 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (-73%), total 1-hydroxypyrene (-53%), urine mutagenicity (-52%), 4-aminobiphenyl hemoglobin adducts (-43%), carboxyhemoglobin AUC7-23 h (-80%), and 3-hydroxypropylmercapturic acid (-35%). These reductions in exposure in the EHCSS group were associated with statistically significant and pathophysiologically favorable changes in several cardiovascular risk factors, including white blood cell count (-0.78 x 10(3)/microL), hemoglobin (-0.16 g/dL), hematocrit (-0.44%), urine 11-dehydrothromboxane B2 (-374 ng/24 h), and high-density lipoprotein cholesterol (+5 mg/dL).

Determination of Environmental Tobacco Smoke from a Second-Generation Electrically Heated Cigarette Smoking System and Conventional Cigarettes

Abstract This substudy of a randomized, controlled, forced switching, open-label, parallel-group clinical study in a total of 100 healthy adult male and female smokers compared environmental tobacco smoke (ETS) produced from smoking a second generation electrically heated cigarette smoking system (EHCSS), two conventional cigarettes, and no-smoking. Concentrations of air constituents including respirable suspended particulate matter (RSP), carbon monoxide (CO), and total volatile organic compounds (TVOCs) and ETS markers including solanesol-related particulate matter (Sol-PM), ultraviolet absorbing particulate matter (UVPM), fluorescing particulate matter (FPM), nicotine and 3-ethenylpyridine (3-EP) were measured in a ventilated, furnished conference room over a period of 2 hours on separate occasions. Except for TVOCs, concentrations of air constituents and ETS markers were reduced by 40% to more than 90% when adult smokers were smoking the EHCSS as compared to smoking conventional cigarettes. CO and most ETS marker concentrations were in the same range as no-smoking.

Reduced Toxicological Activity of Cigarette Smoke by the Addition of Ammonia Magnesium Phosphate to the Paper of an Electrically Heated Cigarette: Subchronic Inhalation Toxicology

Cigarette smoke is a complex chemical mixture that causes a variety of diseases, such as lung cancer. With the electrically heated cigarette smoking system (EHCSS), temperatures are applied to the tobacco below those found in conventional cigarettes, resulting in less combustion, reduced yields of some smoke constituents, and decreased activity in some standard toxicological tests. The first generation of electrically heated cigarettes (EHC) also resulted in increased formaldehyde yields; therefore, a second generation of EHC was developed with ammonium magnesium phosphate (AMP) in the cigarette paper in part to address this increase. The toxicological activity of mainstream smoke from these two generations of EHC and of a conventional reference cigarette was investigated in two studies in rats: a standard 90-day inhalation toxicity study and a 35-day inhalation study focusing on lung inflammation. Many of the typical smoke exposure-related changes were found to be less pronounced after exposure to smoke from the second-generation EHC with AMP than to smoke from the first-generation EHC or the conventional reference cigarette, when compared on a particulate matter or nicotine basis. Differences between the EHC without AMP and the conventional reference cigarette were not as prominent. Overall, AMP incorporated in the EHC cigarette paper reduced the inhalation toxicity of the EHCSS more than expected based on the observed reduction in aldehyde yields.

Short‐Term Clinical Exposure Evaluation of a Second‐Generation Electrically Heated Cigarette Smoking System

This randomized, controlled, forced-switching, open-label, parallel-group study in 100 adult male and female smokers of conventional cigarettes evaluated 8 biomarkers of tobacco smoke exposure. After baseline exposure determinations, adult smokers were switched to a second-generation electrically heated cigarette smoking system (EHCSS) for 8 days in a clinical setting. After 8 days of smoking the EHCSS biomarkers of exposure decreased by 43% to 85% compared to baseline. After correction for residual effects (carryover effects due to long elimination half-life and non-tobacco-confounding sources of exposure), reductions in exposure ranged from 59% to 97%. Results from this short-term clinical exposure study indicate that switching from a conventional cigarette to a second-generation electrically heated cigarette smoking system substantially reduced the exposure to several measured potentially harmful constituents of tobacco smoke.

Effects of Levels of Cigarette Smoke Exposure on Symptom‐Limited Spiroergometry

Previous investigations demonstrated reduced exposure to selected cigarette smoke constituents in adult smokers switching from conventional cigarettes (CC) to an electrically heated cigarette smoking system (EHCSS). This study investigated whether reduced exposure and no smoking (NS) would improve exercise performance. In a 3-period crossover study, 18 male adult smokers (age, 43.6+/-5.3 years) of CC were randomized to smoke CC (tar, 11 mg; nicotine, 0.8 mg; carbon monoxide, 11 mg), to use EHCSS (tar, 3 mg; nicotine, 0.2 mg; carbon monoxide, 0.4 mg [Federal Trade Commission method]), or to NS for 3 days before performing symptom-limited spiroergometry. NS and EHCSS vs CC resulted in less severe dyspnea (NS, 44.4% [P<.01 vs CC;] EHCSS, 50% [P=.03 vs CC;] CC, 88.9%), higher working capacity (NS, 2.92+/-0.4 W/kg [P=.06 vs CC;] ECHSS, 2.92+/-0.4 W/kg [P=.04 vs CC;] CC, 2.86+/-0.5 W/kg), higher peak oxygen uptake (NS, 2694+/-466 mL O(2)/min [P=.08 vs CC;] EHCSS, 2830+/-606 mL O(2)/min [P=.03 vs CC;] CC, 2682+/-492 mL O(2)/min), higher anaerobic threshold (NS, 1324+/-306 mL O(2)/min; EHCSS, 1396+/-312 mL O(2)/min [P=.03 vs CC;] CC, 1315+/-290 mL O(2)/min), and higher maximum rate-pressure product (NS, 30.1+/-2.7 x 10(3) mm Hg/min; EHCSS, 2.8 x 10(3) mm Hg/min [P<.01 vs CC;] CC, 30.7+/-29.2+/-3.6 x 10(3) mm Hg/min) indicating that reduced exposure from tobacco smoke and NS for 3 days may improve cardiovascular function as detected by symptom-limited spiroergometry.

The mouse lymphoma thymidine kinase assay for the assessment and comparison of the mutagenic activity of cigarette mainstream smoke particulate phase

The mouse lymphoma thymidine kinase assay (MLA) has been optimized to quantitatively determine the in vitro mutagenicity of cigarette mainstream smoke particulate phase. To test whether the MLA is able to discriminate between different cigarette types, specially constructed cigarettes each containing a single tobacco type – Bright, Burley, or Oriental – were investigated. The mutagenic activity of the Burley cigarette was statistically significantly lower, up to approximately 40%, than that of the Bright and Oriental cigarettes. To determine the impact of two different sets of smoking conditions, American-blend cigarettes were smoked under US Federal Trade Commission/International Organisation for Standardisation conditions and under Massachusetts Department of Public Health (MDPH) conditions. Conventional cigarettes – eight from the US commercial market plus the Reference Cigarettes 1R4F and 2R4F – and an electrically heated cigarette smoking system (EHCSS) prototype were tested. There were no statistically significant differences between the two sets of smoking conditions on a per mg total particulate matter basis, although there was a consistent trend towards slightly lower mutagenic activity under MDPH conditions. The mutagenic activity of the EHCSS prototype was distinctly lower than that of the conventional cigarettes under both sets of smoking conditions. These results show that the MLA can be used to assess and compare the mutagenic activity of cigarette mainstream smoke particulate phase in the comprehensive toxicological assessment of cigarette smoke.

Short‐Term Exposure Evaluation of Adult Smokers Switching From Conventional to First‐Generation Electrically Heated Cigarettes During Controlled Smoking

This randomized, controlled study in 110 male and female adult smokers evaluated biomarkers of tobacco smoke exposure (carbon monoxide [CO], carboxyhemoglobin [CO-Hb], nicotine, urine mutagenicity) under controlled smoking conditions when adult smokers of 1 conventional cigarette brand (CC1) were switched to an electrically heated cigarette smoking system (EHCSS) or a low-tar conventional cigarette (CC2). Baseline exposure was determined while all subjects smoked CC1. Subjects then were stratified for gender and cigarette consumption and randomized to 1 of 5 groups-EHCSS1, EHCSS2, CC1, CC2, or no smoking-and monitored for 8 days. Compared to baseline, biomarkers of exposure on day 8 decreased 53% to 93% (P < .0001) for EHCSS groups and 18% to 39% (P < .02) for CC2. Environmental tobacco smoke arising from the smoking activities of the different study groups was measured in the air of a separate smoking room over 1-hour periods. Concentrations of respirable suspended particulates in both EHCSS groups were about 90% lower than in the CC1 and CC2 groups, similar to the 95% reduction in the no-smoking group. CO was undetectable in the EHCSS and no-smoking groups. Results from this short-term clinical study indicate that switching from a conventional cigarette to a first-generation EHCSS reduces the generation of environmental tobacco smoke and can reduce the exposure to the measured, potentially harmful constituents in tobacco smoke if smokers do not compensate by numbers of cigarettes. The study design was found to be suitable for the evaluation of the exposure of adult smokers to the measured smoke constituents and to allow the differentiation of different cigarette designs.

Analysis of aromatic amines in cigarette smoke

A method for the analysis of o-toluidine, o-anisidine, 2-naphthylamine, and 4-aminobiphenyl in cigarette mainstream smoke has been developed, which combines the sensitivity of their pentafluoropropionyl (PFP) derivatives in negative ion chemical ionization (NICI) mode with the selectivity of the gas chromatography/tandem mass spectrometry (GC/MS/MS) technique. The use of four deuterated analogues as internal standards along with the application of the standard addition method results in accurate and precise results; the interday precision for the aromatic amines was 3-10% and the accuracy ranged from 97-100%. This method was applied to two American-blend University of Kentucky reference cigarettes, eight American-blend market cigarettes, a bright (flue-cured) tobacco cigarette, and an electrically heated cigarette smoking system (EHCSS). For the American-blend cigarettes there was a linear correlation between aromatic amine yields and mainstream smoke 'tar' ('tar' = total particulate matter - (nicotine + water)), whereas the bright tobacco cigarette and the EHCSS demonstrated significantly lower aromatic amine yields on an equal 'tar' basis. The results support the hypothesis that the nitrogen content of the tobacco, and above all the cigarette combustion temperature, are determining factors for the yields of aromatic amines in smoke.

Micronucleus induction in keratinocytes following in vivo skin application of carcinogens

Alkylating agents occur in the environment and are formed endogenously. Tobacco smoke contains a variety of alkylating agents or precursors including, among others, N-nitrosodimethylamine (NDMA), 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), acrylonitrile and ethylene oxide. We developed and validated a method for the simultaneous determination of methylmercapturic acid (MMA, biomarker for methylating agents such as NDMA and NNK), 2-hydroxyethylmercapturic acid (HEMA, biomarker for ethylene oxide) and 2-cyanoethylmercapturic acid (CEMA, biomarker for acrylonitrile) in human urine using deuterated internal standards of each compound. The method involves liquid/liquid extraction of the urine sample, solid phase extraction on anion exchange cartridges, derivatization with pentafluorobenzyl bromide (PFBBr), liquid/liquid extraction of the reaction mixture and LC–MS/MS analysis with positive electrospray ionization. The method was linear in the ranges of 5.00–600, 1.00–50.0 and 1.50–900 ng/ml for MMA, HEMA and CEMA, respectively. The method was applied to two clinical studies in adult smokers of conventional cigarettes who either continued smoking conventional cigarettes, were switched to test cigarettes consisting of either an electrically heated cigarette smoking system (EHCSS) or having a highly activated carbon granule filter that were shown to have reduced exposure to specific smoke constituents, or stopped smoking. Urinary excretion of MMA was found to be unaffected by switching to the test cigarettes or stop smoking. Urinary HEMA excretion decreased by 46 to 54% after switching to test cigarettes and by approximately 74% when stopping smoking. Urinary CEMA excretion decreased by 74–77% when switching to test cigarettes and by approximately 90% when stopping smoking. This validated method for urinary alkylmercapturic acids is suitable to distinguish differences in exposure not only between smokers and nonsmokers but also between smoking of conventional and the two test cigarettes investigated in this study.