Quantitation of flavor-related alkenylbenzenes in tobacco smoke particulate by selected ion monitoring gas chromatography-mass spectrometry

Little is known about the possible health effects associated with inhaling alkenylbenzenes through cigarette smoking, even though these flavor-related compounds have known toxic effects in animals. We developed a rapid and sensitive solid-phase extraction (SPE) method to quantify seven alkenylbenzenes and piperonal in mainstream cigarette smoke particulate. The smoke particulate fraction of a single cigarette was collected on Cambridge filter pads, solvent extracted, concentrated, purified with SPE, and analyzed by selected ion monitoring gas chromatography-mass spectrometry. We positively identified and quantified five alkenylbenzenes compounds (eugenol, isoeugenol, methyleugenol myristicin, and elemicin) and piperonal in the smoke particulate from eight U.S. brands with mean levels (measured in triplicate) ranging from 6.6 to 4210 ng per cigarette. Additionally, complete blocking of nearly invisible ventilation holes in the cigarette filter increased 2- to 7-fold the percent transfer of alkenylbenzenes from tobacco to the particulate fraction of mainstream smoke.     

Determination of 10 carcinogenic polycyclic aromatic hydrocarbons in mainstream cigarette smoke

Polycyclic aromatic hydrocarbons (PAHs) are one class of chemical compounds that (1) are present at low to trace levels in unburned cigarette filler, and (2) are predominantly generated during combustion. According to a recent report of the International Agency for Research on Cancer, 10 carcinogenic PAHs together with 53 other known carcinogens are present in cigarette smoke. Accurate quantification of these chemicals helps assess public health risk to both smokers and nonsmokers exposed to second-hand smoke. We have developed and validated a specific and sensitive method for measuring these 10 carcinogenic PAHs in the particulate phase of mainstream tobacco smoke. Cigarette smoke particulate, produced using standard machine smoking protocols, was collected on glass fiber Cambridge filter pads. The particulate matter was solvent extracted, purified by solid-phase extraction, and analyzed by liquid chromatography/atmospheric pressure photoionization tandem mass spectrometry using isotopically labeled analogues as internal standards. Our method's limits of detection ranged from 11 to 166 pg and achieved sufficient reproducibility and accuracy to provide useful information on a range of cigarettes having dramatically different machine-smoked tar and nicotine deliveries. The identity of each PAH analyte was established from chromatographic retention time, analyte-specific fragmentation patterns, and relative peak area ratios of the product/precursor ion pairs. This new method provides higher sensitivity, specificity, and throughput than did earlier methods. We found relatively consistent PAH levels among a selection of domestic full-flavor cigarettes. The PAH levels in smoke from highly ventilated light and ultralight cigarettes were low when smoked using ISO (International Organization for Standardization) conditions. However, if highly ventilated cigarettes were smoked under more intense conditions (e.g., larger or more frequent puffs, vents blocked), their PAH levels equaled or exceeded their full-flavor counterparts under ISO conditions.     

Cigarette smoke composition. Part 2. Method for determining major components in smoke of cigarettes that heat instead of burn tobacco

A method is described for determining major constituents in the smoke of a cigarette that heats, but does not burn, tobacco. Dual, simultaneous separations are performed in a single gas chromatographic oven to determine water, glycerol, nicotine, and propylene glycol in a rapid and cost-effective manner. A materials balance of new cigarette smoke total particulate matter was attempted from both Cambridge filter and electrostatic precipitation smoke collection data. Serious deficiencies were found when Cambridge filter smoke collection was applied for this purpose. Electrostatic precipitation smoke collection eliminated these problems. The data obtained by electrostatic precipitation smoke collection indicate that water, glycerol, nicotine, and propylene glycol make up about 94% of new cigarette smoke total particulate matter.     

Cigarette smoke composition. Part 1. Limitations of FTC method when applied to cigarettes that heat instead of burn tobacco

The design of a new cigarette that heats rather than burns tobacco calls for modifications to the Federal Trade Commission (FTC) method for analytical smoking. These changes include eliminating sample conditioning at 75 degrees F and 60% RH, exercising greater care in lighting cigarettes, and smoking cigarettes to self-extinguishment rather than to a predetermined butt length as a measure of complete consumption. By several gross analytical measures, smoke condensate from the new cigarette differs substantially from that of tobacco-burning cigarettes. This is inferred from the lack of coloration of smoke condensate collected on Cambridge filters. Elemental analysis demonstrates reduced carbon and nitrogen content concurrent with increased hydrogen. Thermogravimetric analysis shows almost quantitative weight loss at Tmax = 220 degrees C. Ultraviolet (UV) spectrophotometric analysis shows greatly reduced levels of tobacco-derived smoke components and qualitative differences in chemical entities being measured. By design, the heat required for smoke formation is supplied by a carbon heat source embedded in the cigarette tip. Tobacco contained in the cigarette is not burned and is exposed to temperature less than 300 degrees C. Thus, it is apparent (1) that smoke from the new cigarette contains little or no "tar" as tar is classically defined, and (2) that the FTC method even as modified to account for cigarette design differences is appropriate only for determination of nicotine and carbon monoxide yielded from this cigarette.     

Liquid chromatographic determination of benzo[a]pyrene in total particulate matter of cigarette smoke

The benzo[a]pyrene (BaP) delivery of reference and commercially available tobacco cigarettes, as well as reference and placebo marijuana cigarettes, is determined using a sequential liquid chromatographic/liquid chromatographic procedure. The total particulate matter of sample cigarette smoke is collected using a Cambridge filter pad, which is ultrasonically extracted with acetone. The resulting extract is filtered, then fractionated using semipreparative-scale normal phase liquid chromatography (LC). Quantitative determination is achieved using analytical-scale reverse phase LC equipped with a fluorescence detector. The method is precise (+/- 10-15% relative standard deviation) and yields 85% or better BaP recovery at the ng/cig. level. A single pad may be analyzed in 8 person-hours, while a more typical lot of 12 pads (6 pads each for 2 cigarette brands) may be analyzed in 10 person-days.     

Identification of components responsible for the odor of cigar smoker's breath

Following smoking 1/2 of a cigar, the most odorous cigar tobacco smoke components extracted from the surface of the tongue by nylon-meshed swabs and then extracted from the swab headspace by solid phase microextraction were ethyl pyrrole, 2,3-dimethyl pyrazine, and 2-ethyl pyridine. Similar classes of compounds were identified from the headspace of an aqueous simulated saliva solution treated with cigar smoke. The most odorous compounds were 2,3,5-trimethyl pyridine, 2,5-dimethyl pyrazine, and 2-ethyl-3,5-dimethyl pyridine. Pyridines and pyrazines, the most prominent classes of odorous compounds identified in this experiment, may be generated during cigar pyrrolysis by cleavage of nicotine or by Maillard reaction.     

Characterization of puff-by-puff resolved cigarette mainstream smoke by single photon ionization-time-of-flight mass spectrometry and principal component analysis

Soft single photon ionization-time-of-flight mass spectrometry (SPI-TOFMS) and principal component analysis (PCA) were applied for the characterization and discrimination of the chemical patterns of all individual cigarette puffs from the 2R4F University of Kentucky research reference cigarette. The SPI-TOFMS was connected to a smoking machine, and 10 cigarettes were smoked under defined smoking conditions. A total of 41 detected mass signals could be clearly assigned to smoke constituents (e.g., unsaturated hydrocarbons, aromatic species, sulfurous compounds, and nitrogen-containing substances). For further analysis, the on-line recorded mass signals were added up for each cigarette puff resulting in a single summed mass spectrum for each puff. The so-achieved puff-by-puff resolved yields were additionally normalized by the corresponding total ion signal, which eliminated the influences of varying amounts of burnt tobacco. These values were incorporated into a PCA to find differences and similarities in the chemical patterns of the individual cigarette puffs. In addition, absolute (without normalization) and normalized puff resolved yields were used to clarify occurring trends. Thereby, it was shown that the chemical pattern of the first cigarette puff was very unique, whereby extraordinary high yields of unsaturated hydrocarbons are mainly responsible for this. Depending on the smoking procedure, the chemical pattern of the second puff can also be separated from the first and the third puff. In this case, nitrogen-containing substances play an important role. Puffs three to eight show only small but observable differences. These changes are greater influenced by oxygen-containing and sulfurous smoke constituents. The findings reveal that the overall chemical patterns of machine-smoked cigarette puffs vary quite a lot during the smoking process. This lets us assume that the burden of hazardous compounds for the human smoker also differs from puff to puff.     

Artifact formation during smoke trapping: an improved method for determination of N-nitrosamines in cigarette smoke

Studies in our laboratory revealed artifactual formation of N-nitrosamines during trapping of mainstream and sidestream tobacco smoke by the method of Hoffmann and coworkers (2, 4). Both volatile and tobacco-specific N-nitrosamines were produced. This artifact formation took place on the Cambridge filter, which is part of the collection train used in the previously published procedure. When the filter was treated with ascorbic acid before smoke collection, artifact formation was inhibited. The improved method resulting from these studies was applied to a comparative analysis of N-nitrosamines in smoke from cigarettes that heat, but do not burn, tobacco (the test cigarette) and several reference cigarettes. Concentrations of volatile and tobacco-specific N-nitrosamines in both mainstream and sidestream smoke from the test cigarette were substantially lower than in the reference cigarettes.     

The form of nicotine in tobacco. Thermal transfer of nicotine and nicotine acid salts to nicotine in the gas phase

Thermal transfer to nicotine in the gas phase from neat nicotine, from various nicotine carboxylic acid salts, and from endogenous nicotine in Burley, Bright, and Oriental tobacco samples has been examined by thermogravimetric/differential thermal analysis/mass spectroscopy and evolved gas analysis. Under the conditions used in these studies, the peak transfer temperatures of these substances to nicotine in the gas phase are nicotine and nicotine acetate, both ca. 110-125 degrees C; nicotine malates, ca. 110-210 degrees C for nicotine to malic acid ratios of 1:0.56 and 1:1 and ca. 160-210 degrees C for a nicotine to malic acid ratio of 1:2; (S)-nicotine bis[(2R,3R)-hydrogen tartrate] dihydrate, ca. 195-210 degrees C; and tobacco samples, a range of ca. 160-220 degrees C. These results suggest that nicotine is mostly protonated in tobacco leaf. In all cases, the temperature of the transfer of nicotine to the gas phase was found to be many hundreds of degrees below the temperatures observed around the coal of a burning cigarette (smolder, ca. 500-775 degrees C; dynamic smoking, 600 to over 950 degrees C). Within the narrow zone of a puffing cigarette that encompasses an intermediate temperature range (125-250 degrees C), kinetic data suggest that these temperatures are not sufficient to volatilize significant amounts of nonprotonated nicotine, assuming any exists at all, during the short puff duration (2 s). It is concluded that nonprotonated nicotine and protonated nicotine (salts of nicotine with natural tobacco carboxylic acids) will transfer nicotine to smoke with comparable yields and efficiencies during the smoking process.     

辐照处理对货架期卷烟品质及微生物数量的影响

为考察辐照对卷烟品质及微生物数量的影响,采用0、4和8kGy的60Coγ射线辐照卷烟,检测样品中霉菌数、酶的活性和霉变现象,在贮存0、180和360d时检测样品的菌落总数、水分、主流烟气焦油释放量,并进行了感官评价试验。结果表明,采用4和8kGy剂量辐照卷烟,能有效杀灭霉菌,防治卷烟霉变;辐照能降低货架期卷烟中的菌落总数、抑制α-淀粉酶和果胶酶的活力,减缓货架期卷烟感官质量下降,其中,4kGy剂量辐照卷烟在货架期的感官质量比对照稍好;辐照对卷烟的焦油和烟碱释放量及水分含量没有明显影响。因此60Coγ射线辐照是一种保持和改善货架期卷烟质量的有效方法。     

Gas chromatographic determination of nicotine in environmental tobacco smoke: collaborative study

A gas chromatographic method for determination of vapor phase nicotine in environmental tobacco smoke (ETS) was collaboratively studied by 6 laboratories. Nicotine is desorbed from XAD-4 sample tubes with ethyl acetate containing triethylamine and determined by gas chromatography with nitrogen-selective detection. Each collaborator received blind duplicate samples at each of 6 nicotine concentrations. Three concentrations were generated by spiking XAD-4 tubes with known amounts of nicotine; the remaining 3 concentrations were ETS samples obtained in a carefully controlled environmental chamber containing sidestream and exhaled mainstream smoke from 1R4F Kentucky reference cigarettes. Repeatability and reproducibility relative standard deviations ranged from 4.4 to 11.1% and from 7.0 to 11.1%, respectively, for nicotine concentrations evaluated (up to 6 micrograms/cu m). The method has been adopted official first action.     

苹果酸脲结晶肥对原烟中氨基酸含量及原烟质量的影响

进行了苹果酸脲结晶肥 (简称结晶肥 )对原烟中游离氨基酸、总糖、烟碱等主要化学成分含量及对原烟质量影响的研究。连续 2年的结果表明 ,原烟中酸性氨基酸、除精氨酸外的碱性氨基酸及多种中性氨基酸含量均随结晶肥施用量的增加而增加 ;结晶肥使原烟中蛋氨酸、鸟氨酸、色氨酸含量明显低于不施肥对照和施等氮量烟草专用复肥的对照处理。随结晶肥用量增加 ,r 氨基丁酸、牛磺酸、亮氨酸、缬氨酸的含量下降 ;精氨酸、胱氨酸、异亮氨酸含量较少。等N量情况下 ,结晶肥增加原烟中K含量 15.6%～19.5% ,增加总糖含量 13.33%～17.70% ,增加还原糖含量 10.5% ,原烟中Cl含量降低 37.5%和烟气中焦油含量降低 16.9% ,达到显著水准。结晶肥使原烟中糖 /碱比例更合理 ,使原烟的评吸质量比复肥对照处理上升 2个等级 ,香气量和浓度各提高 1个等级 ,评吸质量档次高 2个等级 ,原烟质量明显改善     

Development of machine smoking parameters for measurement of cigarette tar yield in the United Kingdom

The smoking and analytical methods used by government and tobacco industry laboratories are the result of many years of collaborative work within the industry and through international bodies such as the International Standards Organization. Recently, some publications have criticized the validity and scientific soundness of these procedures. It is shown that such criticisms are totally unfounded by reviewing the work that led to the adoption of the current technique. Tar tables, published by the UK Health Departments are valid in ranking brands in order of their yields, while not necessarily reflecting the absolute tar delivery to the smoker. These tables are intended to and do provide a relevant guide for smokers about the proportional tar yields of UK cigarettes.     

Chemical characterization of dissolvable tobacco products promoted to reduce harm

In 2009, the R. J. Reynolds Tobacco Co. released a line of dissolvable tobacco products that are marketed as an alternative to smoking in places where smoking is prohibited. These products are currently available in Indianapolis, IN, Columbus, OH, and Portland, OR. This paper describes the chemical characterization of four such products by gas chromatography-mass spectrometry (GC-MS). The dissolvable tobacco products were extracted and prepared by ultrasonic extraction using acetone, trimethylsilyl derivatization, and headspace solid phase microextraction (SPME). The following compounds were identified in the dissolvables using either ultrasonic extractions or trimethylsilyl derivatization: nicotine, ethyl citrate, palmitic acid, stearic acid, sorbitol, glycerol, and xylitol. The following compounds were identified in the dissolvables using headspace SPME: nicotine, ethyl citrate, cinnamaldehyde, coumarin, vanillin, and carvone. With the exception of nicotine, the compounds identified thus far in the dissolvables are either flavoring compounds or binders. The concentration of free nicotine in the dissolvables was determined from the Henderson-Hasselbalch equation and by measuring the pH and nicotine concentration by GC-MS. The results presented here are the first to reveal the complexity of dissolvable tobacco products and may be used to assess potential oral health effects.     

Quantification of flavor-related compounds in the unburned contents of bidi and clove cigarettes

Bidi cigarettes, small hand-rolled cigarettes produced primarily in India, are sold in the United States in a wide variety of candy-like flavors (e.g. dewberry, chocolate, clove) and are popular with adolescents. Many flavored bidis contain high concentrations of compounds such as eugenol, anethole, methyleugenol, pulegone, and estragole; several of these compounds have known toxic or carcinogenic properties. Clove cigarettes, or kreteks, are another highly flavored tobacco product with high levels of eugenol due to clove buds present in the tobacco filler. In this study, compounds in the burnable portion-the filler and wrapper material actually consumed during the smoking of bidis, kreteks, and U.S. cigarettes-were analyzed. Flavor-related compounds were solvent extracted from the burnable portion of each cigarette with methanol. An aliquot of the methanol extract was heated, and the sample headspace was sampled with a solid-phase microextraction fiber and introduced into a gas chromatograph-mass spectrometer for analysis in selected-ion monitoring mode. High levels of eugenol were detected in five clove-flavored bidi brands ranging from 78.6 to 7130 microg/cigarette (microg/cig), whereas diphenyl ether (128-3550 microg/cig) and methyl anthranilate (154-2360 microg/cig) were found in one grape-flavored bidi brand. A nontobacco herbal bidi brand contained the greatest variety of compounds, including anethole (489-665 microg/cig), eugenol (1670-2470 microg/cig), methyleugenol (27.7-36.6 microg/cig), safrole (32.4-34.4 microg/cig), myristicin (170-247 microg/cig), and elemicin (101-109 microg/cig). Filler from kreteks was found to contain high levels of eugenol, anethole, and coumarin. Flavored bidis and clove cigarettes contain a number of compounds that are present at levels far exceeding those reported in U.S. cigarette tobacco. Research is underway to determine the levels of these compounds delivered in smoke. It is not known what effect inhalation of these compounds has on smokers.     

烤烟烟碱含量与土壤有机质、氮素含量的关系分析

分析了湖南烟区222个烤烟样品烟碱含量与土壤有机质、全氮和碱解氮之间的关系。结果表明：（1）烤烟烟碱含量在不同等级间存在较大的变异，总体呈现出B2F〉G3F〉X2F的趋势，平均烟碱含量为2．84％，变幅为0．58％～5．95％，变异系数为34．07％；（2）不同等级烟碱含量分别与土壤有机质、全氮和碱解氮的相关系数均达到1％的极显著水平；（3）采用聚类分析方法把同一等级烟叶样品的烟碱含量分为高、中、低3类，不同类别相比较，有机质含量越高的土壤，其土壤全氮、碱解氮以及相应的烟叶烟碱含量也越高，表明土壤有机质含量的高低，直接影响了土壤氮素的供应状况，进而影响了烟碱含量的积累。     

烟叶和卷烟辐照醇化效果的研究

实验结果表明９ｋＧｙ辐照初烤原烟和机烤烟叶醇化效果较明显，可以缩短烟叶自然醇化期１～１．５ａ，７～９ｋＧｙ辐照杀菌效果明显，对未加香烟丝和成品卷烟的醇化效果也较明显，５～９ｋＧｙ辐照，既不能降低焦油含量，也不能产生诱发放射性，在５～１００ｋＧｙ范围内，烟叶和卷烟的化学成分变化不明显。     

能量密度对低温等离子体辅助催化转化柴油机颗粒物和NOx的影响

为了研究低温等离子体辅助催化后处理技术对柴油机有害排放物的作用效果,基于介质阻挡放电理论设计了低温等离子体（NTP）发生器,研究了NTP发生器工作电压随能量密度的变化关系。以涂覆γ-Al2O3的蜂窝状陶瓷为载体,采用柠檬酸凝胶法制备了LaMnO3催化剂,建立了NTP辅助LaMnO3催化试验系统。在台架试验基础上,研究了能量密度变化对NTP辅助LaMnO3催化技术转化（颗粒物）PM和NOx的影响规律。试验结果表明：在相同频率作用下,NTP发生器的能量密度随着工作电压的升高而增加;与原机相比,NTP辅助LaMnO3催化技术作用后NO浓度显著降低,NO2浓度显著增加,NOx浓度降低;随着能量密度的增加,碳烟转化效果越明显,碳烟的不透光烟度最高降低约83%。结果表明NTP辅助LaMnO3催化技术可有效转化PM和NOx,在柴油机排气后处理领域有着广泛的研究前景。     

Gender, environmental tobacco smoke, and pulmonary function in rural children and adolescents: the Humboldt study

We examined the gender-related association between household exposure to environmental tobacco smoke (ETS) and pulmonary function among 862 children and adolescents aged 6 to 17 years living in the town of Humboldt, Saskatchewan, in 1993. Pulmonary function tests included forced vital capacity (FVC), forced expiratory volume in one second (FEV1), maximum mid-expiratory flow rate (FEF25-75), and flow rates at 75%, 50%, and 25% of vital capacity (Vmax75, Vmax50, and Vmax25). Each pulmonary function test variable was regressed on age, height, weight, and their quadratic and cubic polynomials, with the terms significant at the alpha level of 0.10 being retained. Residuals for the pulmonary function test variables, which are the differences between the observed and predicted values, were calculated. Estimations of ETS exposure were parental smoking status, number of household smokers, total daily cigarette consumption, and number of cigarettes smoked daily at home by household members. Maternal smoking status was significantly related to residual FEF25-75, Vmax75, Vmax50, and Vmax25. Number of household smokers and daily cigarette consumption by household members were significantly associated with FEV1, FEF25-75, Vmax75, Vmax50, and Vmax25, and the association between ETS exposure and pulmonary function was stronger in girls than in boys. Interaction of gender and number of cigarettes smoked daily at home was significantly related to FEF25-75, Vmax75, Vmax50, and Vmax25 among the non-smoking subjects. We concluded that ETS exposure had a larger effect on pulmonary function in girls than in boys.     

Evaluation of two derivatization reagents for the determination by LC-MS/MS of ammonia in cigarette mainstream smoke

Ammonia in cigarette mainstream smoke was quantified by LC-MS/MS after derivatization. Two different reagents, fluorescamine and dansyl chloride, were investigated, but only the latter gave stable derivatives; therefore, it was considered the most appropriate choice. Smoke samples were collected on a Cambridge filter pad followed by an impinger containing a solution of hydrochloric acid. Ammonia was then derivatized with a 18.5 mM solution of dansyl chloride in acetonitrile at 70 °C for 30 min in a vial with the internal standard, (15)ND(4)Cl. The resulting derivative was analyzed by LC-MS/MS detection with an atmospheric pressure chemical ionization (APCI) interface in the positive ionization mode using multiple-reaction monitoring (MRM). Good linearity was obtained in the concentration range of 0.02-1.65 μg/mL (r(2) ≥ 0.999), and the limit of detection (LOD) was established at 0.006 μg/mL. This method has the advantage of being sensitive, efficient, and reliable and is not hindered by interferences from the sample matrix. It should thus be considered a reference method of choice for the determination of ammonia in smoke.