The vapor pressure of nicotine has been reported for unprotonated nicotine and for nicotine-water solutions. Yet no published values exist for nicotine in any commercially relevant matrix or for protonated forms (e.g., tobacco, smoke, electronic cigarette solutions, nicotine replacement products, nicotine salts). Therefore a methodology was developed to measure nicotine activity (defined as the vapor pressure from a matrix divided by the vapor pressure of pure nicotine). The headspace concentration of nicotine was measured for pure nicotine and tobacco stored at 23, 30, and 40 °C which allowed for conversion to vapor pressure and nicotine activity and for the estimation of enthalpy of vaporization. Burley, Flue-cured, Oriental, and cigarette blends were tested. Experiments were conducted with pure nicotine initially until the storage and sampling techniques were validated by comparison with previously published values. We found that the nicotine activity from tobacco was less than 1% with Burley > Flue-cured > Oriental. At 23 °C the nicotine vapor pressure averaged by tobacco type was 0.45 mPa for Oriental tobacco, 1.8 mPa for Flue-cured, 13 mPa for Burley while pure nicotine was 2.95 Pa. In general, the nicotine activity increased as the (calculated) unprotonated nicotine concentration increased. The nicotine enthalpy of vaporization from tobacco ranged from 77 kJ/mol to 92 kJ/mol with no obvious trends with regard to tobacco origin, type, stalk position or even the wide range of nicotine activity. The mean value for all tobacco types was 86.7 kJ/mol with a relative standard deviation of 6.5% indicating that this was an intrinsic property of the nicotine form in tobacco rather than the specific tobacco properties. This value was about 30 kJ/mol greater than that of pure nicotine and is similar to the energy needed to remove a proton from monoprotonated nicotine.
The present study describes the development of a liquid chromatography tandem mass spectrometry (LC-MS/MS) technique for the analysis of trace levels of four tobaccospecific nitrosamines (TSNAs): nitrosoanabasine (NAB), nitrosoanatabine (NAT), 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), and nitrosonornicotine (NNN). The technique can be applied for the analysis of TSNAs in USP grade nicotine. Nicotine used in e-liquids for the electronic smoking devices is typically obtained from tobacco plant materials (Nicotiana tabacum, Nicotiana rustica) and, although it is purified, it contains besides nicotine low levels of several contaminants such as minor alkaloids. It also contains traces of TSNAs. Analysis of TSNAs in USP grade nicotine is a challenging task since the analyzed samples contain about 10+7–10+8 times more nicotine than individual TSNAs. Because the analyzed solutions cannot be diluted too much in order to keep the TSNAs level above the limit of quantitation (LOQ), even for apparently good chromatographic separations, the peak tailing of nicotine may generate interferences. The new method of analysis uses a Luna Omega 1.6 μm particles chromatographic column for separation and detection on a LC-MS/MS instrument with scheduled multiple reaction monitoring (Scheduled MRM). The levels of TSNAs in nicotine of USP purity from four commercial sources varied between 3 to 8 ng/g NAB, 4 to 20 ng/g NAT, 30 to 50 ng/g NNK, and 0.5 to 2 ng/g for NNN. Besides the analysis of TSNAs in nicotine, the technique has been applied successfully in the analysis of TSNAs in e-liquids and in particulate phase generated by the electronic smoking devices.
The present study evaluated the in vitro extraction of benzo[a]pyrene (BaP) from moist snuff into water and into artificial saliva. A similar, previous study evaluated the levels of BaP that remained in the moist snuff after the extraction but did not measure the levels of BaP in the water or saliva extract. The previous study showed that the remaining levels of BaP in the solid material were between 96.3% and 109.6% relative to the initial level of BaP, when the snuff was washed with water and between 99.4% and 108.3% from the initial level of BaP, when the snuff was washed with both saliva and water. Nine moist snuff samples (eight being the same brands as evaluated in the previous study) were analyzed in the present study. Several improvements were made compared to the previous study regarding the extraction conditions. The extraction was performed for 1 h at 37 °C, using a mechanical agitator.
The previous study used a commercially available artificial saliva which had an adjusted pH but did not contain enzymes or salts. This saliva was replaced with complete artificial saliva containing salts, mucin and enzymes. The results indicated that the level of BaP extracted in 100 mL water from 5 g of moist snuff at 37 °C ranged between 1.0% and 1.7% of the initial level present in tobacco. For artificial saliva, the extracted level of BaP was between 2% and 3.9% from the initial level, depending on the moist snuff brand. Although the BaP level extracted from the moist snuff with artificial saliva remained very low, the surfactant character of artificial saliva increased BaP extraction relative to water by a factor of approximately two. This study supports the previous reported finding that the vast majority of BaP in moist snuff is not extracted in water or artificial saliva. [Beitr. Tabakforsch. Int. 29 (2020) 21–26]
The relatively volatile nature of the particulate matter fraction of e-cigarette aerosols presents an experimental challenge with regard to particle size distribution measure-ments. This is particularly true for instruments requiring a high degree of aerosol dilution. This was illustrated in a previous study, where average particle diameters in the 10-50 nm range were determined by a high-dilution, electrical mobility method. Total particulate matter (TPM) masses calculated based on those diameters were orders of magnitude smaller than gravimetrically determined TPM. This discrepancy was believed to result from almost complete particle evaporation at the dilution levels of the electrical mobility analysis. The same study described a spectral transmission measurement of e-cigarette particle size in an undiluted state, and reported particles from 210-380 nm count median diameter. Observed particle number concentrations were in the 109 particles/cm3 range. Additional particle size measurements described here also found e-cigarette particle size to be in the 260-320 nm count median diameter range. Cambridge filter pads have been used for decades to determine TPM yields of tobacco burning cigarettes, and collection of e-cigarette TPM by fibrous filters is predicted to be a highly efficient process over a wide range of filtration flow rates. The results presented in this work provide support for this hypothesis.
Described here is a study in which e-cigarette aerosols were collected on Cambridge filters with adsorbent traps placed downstream in an effort to capture any material passing through the filter. Amounts of glycerin, propylene glycol, nicotine, and water were quantified on the filter and downstream trap. Glycerin, propylene glycol, and nicotine were effciently captured (> 98%) by the upstream Cambridge filter, and a correlation was observed between filtration efficiency and the partial vapor pressure of each component. The present analysis was largely inconclusive with regard to filter efficiency and particle-vapor partitioning of water. [Beitr. Tabakforsch. Int. 26 (2014) 183-190]
The present study describes the analysis of several organic acids in tobacco and smokeless tobacco products using a liquid chromatography (LC) method with mass spectrometric (MS) detection (LC-MS). Prior to the application of the LC-MS method, a qualitative analysis for the identification of the organic acids in tobacco and oral tobacco products was performed. The qualitative method used direct silylation of the plant material followed by GC-MS separation and detection. For the acids’ quantitation, a novel LC-MS method has been developed and validated. The acids of interest for quantitation were the following: acetic, citric, fumaric, glyceric, lactic, maleic, malic, oxalic, pyroglutamic, pyruvic, quinic, and trihydroxybutanoic. The LC separation was performed on a Synergy 4u Hydro-RP column 250 × 4.6 mm, with an aqueous mobile phase containing 5% methanol and 0.15% formic acid. The LCMS method has the advantage versus LC methods with other detection types (refractive index, UV absorption at low UV range, or conductivity) of being capable of positive identification of the analytes based on their specific ions, and of having significantly better sensitivity. Unfortunately, the LC-MS method was not generating good results for oxalic acid and acetic acid also expected to be present in some samples of tobacco or tobacco products. The study describes the advantages and disadvantages of the LC-MS method for the analysis of organic acids in tobacco and smokeless tobacco products.
The present study evaluated in vitro extractability of various polycyclic aromatic hydrocarbons (PAHs) from moist snuff, when the extracting agent was water or artificial saliva. The extraction was performed on nine brands of moist snuff samples that are commercially available and were purchased from the market in January 2018. The moist snuff brands were selected to represent brands with different tobacco cut size descriptors and flavors. For the measurement of PAHs, two different analytical methods were used, an HPLC (High Performance Liquid Chromatography) method for measuring only benzo[a]pyrene (BaP) and a GC/MS/MS (Gas Chromatography Tandem Mass Spectrometry) method for measuring 21 PAHs (including BaP). These methods were modifications of preexistent methods reported in the literature. The results for BaP indicated that by extracting 500 mg of freeze-dried moist snuff with 6 portions of 20 mL water (120 mL), or with 4 portions of 20 mL artificial saliva, followed by two portions of 20 mL water, the BaP remains close to 100% in the solid material and it is not detected in the extracting solution. PAHs with a molecular weight similar or heavier than BaP also showed no extractability. Lighter PAHs such as fluorene, phenanthrene, anthracene, and 5-methylanthracene showed a relatively good extractability. An intermediate group including fluoranthene, pyrene, and benz[a]anthracene showed some extractability in the conditions of this in vitro experiment. This study is not a substitute for clinical studies regarding PAH uptake in human users of moist snuff. However, the results indicate very limited bioavailability of BaP and heavier PAHs from moist snuff. Higher, but variable bioavailability was indicated for lighter PAHs. Important implications of these findings are that: 1) measurably different BaP content of two moist snuff products is unlikely to result in any meaningfully different consumer exposure to BaP; and 2) biomarkers for one PAH cannot necessarily be used as a reliable indicator of exposure to another PAH, particularly if the molecular weights of the precursor PAHs differ since their bioavailabilities can be very different. [Beitr. Tabakforsch. Int. 28 (2019) 214–223]
A reliable and sensitive method for the measurement of the level of diacetyl (2,3-butanedione) and acetylpropionyl (2,3-pentanedione) in the aerosol (both the particles and the suspending gas) of electronic smoking devices (e-cigarettes) has been developed. The method uses a gas chromatographic separation on a Carbowax type column with the measurement of the analytes on a triplequadrupole mass spectrometer working in positive MRM mode. The method has been validated using standard requirements regarding selectivity, sensitivity, recovery, accuracy, and repeatability. The limit of quantitation (LOQ) for the method was determined to be 0.41 ng/mL for diacetyl and 0.21 ng/mL for acetylpropionyl as measured for standards. These values translate to an LOQ of 0.082 ng/puff for diacetyl and 0.042 ng/puff for acetylpropionyl as measured for an e-cigarette with 50 puffs placed in 10 mL acetone. The samples analyzed included collected aerosols from several e-cigarettes, and a number of liquids used in electronic cigarettes (e-liquids). 3R4F Kentucky reference cigarette was also analyzed for evaluating the accuracy of the procedure, with good agreement with data from the literature. Diacetyl and acetylpropionyl were distributed in both particulate phase and also in vapor phase. The levels of diacetyl and acetylpropionyl in particulate phase collected from 3R4F cigarettes were found to represent only about 22% for diacetyl and only 31% for acetylpropionyl, while the vapor phase for diacetyl represented 78% and for acetylpropionyl 69% of the total analyte. The levels of diacetyl and acetylpropionyl in the aerosols of most electronic smoking devices were found to be very low, with a few exceptions. The analysis of the two analytes in several e-liquids available on the market showed a very large range of levels. Some of the e-liquids from the market are likely to have diacetyl and/or acetylpropionyl intentionally added.
Present study describes the determination of nicotine in various plant samples with a low content of this compound. Nicotine is found naturally in plants from the Solanaceae family. The plants from Nicotiana genus contain large levels of nicotine. However, only low levels are present in plants from Solanum genus including potato, tomato, eggplant, and from Capsicum genus, which are used as food. Because the levels of nicotine in these materials are in the range of parts per billion, the measurements are difficult and the results are very different from study to study. The present study evaluated the level of nicotine in a number of plants (fruits, roots, leaves, tubers) from Solanaceae family (not including Nicotiana genus) and from several other vegetables commonly used as food. The analysis consisted of the treatment of plant material with an aqueous solution 5% NaOH at 70°C for 30 min, followed by extraction with TBME containing d3-nicotine as an internal standard. The TBME organic layer was analyzed on a 7890B/7000C GC-MS/MS system with a 30 m × 0.25 mm, 0.25 μm film CAM column. The MS/MS system worked in MRM positive ionization mode monitoring the transition 162 - 84 for nicotine and 165 - 87 for d3-nicotine. Particular attention was given to the preservation of the intact levels of nicotine in the plant material. The plant material was analyzed as is, without drying and with minimal exposure to contaminations. Separately, the moisture of the plant material was measured in order to report the nicotine level on a dry-basis. Levels of nicotine around 180 ng/g dry material were obtained for tomatoes and eggplant (fruit) and lower levels were obtained for green pepper and potato. Similar levels to that in the tomato fruit were detected in tomato leaves. Materials from other plant families also showed traces of nicotine. [Beitr. Tabakforsch. Int. 27 (2016) 54-59]
The present study describes a reliable technique for the analysis of free amino acids in tobacco leaf. The levels of amino acids in tobacco are important since they are related to both tobacco quality and the potential generation in cigarette smoke of toxicants having amino acid precursors. Other techniques used in the past for amino acid analysis have various shortcomings that were avoided in the present method. The new method uses HPLC separation and a tandem mass spectrometer for detection with no derivatization step as sample preparation. The separation has been obtained using ion pair HPLC on a reversed phase column that offers excellent chromatographic resolution. The MS/MS detection procedure offers very good sensitivity and positive identification of the analytes. The procedure was fully validated and can be used for the analysis of 24 amino acids. It was applied for the quantitation of amino acids from 16 types of tobacco including flue-cured and Burley, some domestic and some not grown in the USA, two types of Oriental tobacco, and from tobacco of a 3R4F Kentucky reference and a common commercial cigarette. It was shown that the analysis provides useful information regarding the amino acid level variation between tobacco types, between tobacco stalk positions, and between the growing locations of different tobaccos. [Beitr. Tabakforsch. Int. 26 (2015) 334-343]