SUMMARY The airtightness of heated tobacco product (HTP) packs is a very important indicator for the product quality and is also of great importance during the conditioning process. A method for evaluation of the airtightness was developed based on the air pressure difference in a constant pumping configuration. The essential feature of this method is that the pressure difference between the inside and the outside of the HTP packs during the deflation process is used to characterize the sealing quality of HTP packs. The detailed setup, the principle as well as the determination procedure are described. The accuracy and the repeatability of the method were assessed, and the effect of airtightness on the conditioning process was also investigated. The developed method is proven to be reliable with a standard deviation less than 0.09 kPa and repeatability less than 0.30 kPa. In addition, it was found that, although the transmission of moisture between HTPs and atmosphere could not be entirely prevented by the packs, airtightness still plays a significant role during the conditioning process, especially if the airtightness was at a relatively low level (e.g., lower than 1.5 kPa under a pumping flow rate of 200 mL/min). The method provides a promising way to assess and monitor the sealing quality of HTP packs, and it is suggested that the airtightness of the pack should not be lower than 2 kPa under a pumping flow rate of 200 mL/min. [Contrib. Tob. Nicotine Res. 32 (2023) 140–145]

Summary High lignin content of tobacco stem has been addressed as a drawback for its utilization on manufacture of reconstituted tobacco sheet. Therefore, the solvothermal method using ethylene glycol was investigated for the removal of lignin from tobacco stem. It was found that the removal efficiency of ethylene glycol on tobacco stem is much lower than that achieved on wheat straw and corncob, i.e., 13.9% vs. 39.3% and 44.1%, respectively. This can be rationalized in terms of the presence of solanesol in tobacco stem, which retards the strong hydrogen bond interaction between ethylene glycol and the free hydroxyl groups present in lignin. When solanesol was eliminated by applying an n-hexane extraction procedure, 40.5% of lignin was successfully dissolved from tobacco stem. The successful removal of lignin was further confirmed by the characterizing results of SEM, FT-IR and N2-sorption. The developed method provides a promising way to attenuate the negative effects of lignin on utilization of the otherwise wasted tobacco stem for production of reconstituted tobacco sheet.

Exogenous dietary advanced glycation endproducts (AGEs) have been suggested to increase health risk. Oral use tobacco products are used as substitute for cigarette smoking due to their "reduced risk" and no second-hand smoke. They may increase AGEs intake due to their oral form of use. However, research on AGEs in oral use tobacco products is limited. A HPLC-MS/MS method was developed to survey free and total N epsilon-Carboxymethyllysine (CML) and N epsilon-Carboxyethyllysine (CEL) levels in different oral use tobacco products in this work. HPLC conditions, such as HPLC column and mobile phase additive were optimized. With the optimized conditions, analytical method performance for three types of oral use tobacco matrix (i.e. CRP2, gum and dissolvable tobacco product) was characterized. CML and CEL showed higher recovery in all three sample matrix, and the recovery of CML and CEL was in the range of 88.7%-120%, 90.0%-110%, respectively. The developed method showed good intra-day and inter-day precision with RSDs for both free and total CML (or CEL) of less than 9.7%. Limit of detections (LODs) of CML and CEL were 0.36 and 0.25 ng/mL, respectively. Sample analysis for 21 oral use tobacco products was conducted with the developed method. As for oral use tobacco products, total CML and CEL was much more than those of their free state, indicating that CML and CEL exist mainly protein adducts. In addition, the content of CML was higher than that of CEL for both free and total state. Oral use tobacco products I (i.e. dry snuff, moist snuff, suns and chew) had the highest CML (i.e. 44.89-608.69 mu g/g) and CEL (i.e. 15.98-77.70 mu g/g) levels, compared with oral use tobacco products II (i.e. gum) with the least CML (i.e. 0.7-0.8 mu g/g) and CEL (i.e. 0.14-0.18 mu g/g) levels. It mainly depended on the composition and processing technology of oral use tobacco products. Both total CML and CEL showed linear relationship with lysine and total amino acid (i. e. R 0.77). However, there was no linear relationship between total CML (or CEL) and total reducing carbohydrates (i.e. R < 0.5).

建立了同时测定人尿液中烟碱及其9种代谢物的高效液相亲水作用色谱-串联质谱分析方法.尿液样品经乙腈和甲醇混合溶剂(体积比3:1)稀释20倍,过0.22μm滤膜后直接进样.采用Atlantis HILIC Silica柱(3.0 mm×100 mm,3.0μm),以10 mmol/L甲酸铵水溶液(用甲酸调至pH 3.0)-乙腈为流动相进行梯度分离,电喷雾正离子电离模式和多反应监测扫描模式下测定,以同位素内标法定量.结果表明,烟碱及其9种代谢物在各自的质量浓度范围内线性关系良好,相关系数(r)均大于0.997,检出限(LOD)和定量下限(LOQ)分别为0.03～0.24 ng/mL和0.10～0.80 ng/mL;在低、中和高3个加标水平下的加标回收率为81.9％～110％,日间和日内相对标准偏差(RSD)为0.50％～6.7％.该方法操作简单、灵敏度高、稳定性好,能够满足大批量人尿液中烟碱及其9种代谢物的检测需求.

本文针对ISO 3402:1999(GB/T 16447-2004)中规定的强制气流对卷烟进行调节平衡进行了研究,平衡效果采用标准规定的相对重量变化率进行判定.结果表明:施加额外的紊乱强制气流可以加快高含水率和低含水率卷烟样品平衡的速率,但对于常规卷烟样品并无明显效果.当调节时间满足标准规定的时间(48h),无论是否施加强制气流,卷烟样品都能达到标准规定的平衡要求.该研究结果对于相关标准的修订以及企业和研究人员对于标准的使用都具有重要的指导意义.

Summary The width of cut tobacco strands is an important indicator for physical parameters as well as for the smoking quality. In some countries, cut width helps to distinguish fine-cut tobacco and pipe tobacco and thus differentiates taxation rate. A new method for rapid measurement of the width of cut tobacco strands was developed based on digital image processing, because the method described in ISO 20193, though easy to implement in factories, proved time consuming and generated high testing costs. The essence of this method is to determine the statistic width of incisions. The straight-line segments represent the width of strands of cut tobacco, from which the determination of the width for randomly placed tobacco strands could be achieved. Five kinds of samples (‘ISO collaborative study samples 0.4 mm, 1.0 mm, 1.6 mm and 3.0 mm’ and ‘Guangdong baked 0.9 mm’) were used to study the comparability of the measurement results between the method presented in this work and the current ISO method. Results show that accuracy and repeatability are comparable. In addition, the testing efficiency of the method presented in this work appears to be higher than the current ISO method, and it is thus a promising alternative method for measuring the width of strands of cut tobacco.

Summary Conditioning is a very important procedure for tobacco and tobacco products before their corresponding chemical and physical analysis. For cigarettes, forced air flow is generally required during the conditioning procedure. A special wind tunnel was designed to investigate how the forced air flow affects the conditioning of cigarettes in a constant climate laboratory. Two types of cigarettes with blended (including flue-cured and burley tobacco strands) and pure flue-cured tobacco strands were selected as test samples. It was found that the conditioning time to achieve the equilibrium could be shortened from 23 h without forced air flow to 7 h with a forced air flow rate of 2m/s. This is mainly due to the exchange of water molecules between cigarette samples and atmosphere being accelerated by applying the forced air flow. It was concluded that a 48 h conditioning period using the described forced air flow rate setup was unnecessary to attain the equilibrium for cigarettes. [Beitr. Tabakforsch. Int. 28 (2019) 224–229]

Glycosides are a kind of highly important natural aromatic precursors in tobacco leaves. In this study, a novel HKUST-1-coated monolith dip-it sampler was designed for the fast and sensitive analysis of trace glycosides using direct analysis in real-time mass spectrometry. This device was prepared in two steps: in situ polymerization of monolith in a glass capillary of dip-it and layer-by-layer growth of HKUST-1 on the surface of monolith. Sufficient extraction was realized by immersing the tip to solution and in situ desorption was carried out by plasma direct analysis in real time. Compared with traditional solid-phase microextraction protocols, sample desorption was not needed anymore, and only extraction conditions were needed to be optimized in this method, including the gas temperature of direct analysis in real time, extraction time, and CH3 COONH4 additive concentration. This method enabled the simultaneous detection of six kinds of glycosides with the limits of detection of 0.02-0.05 μg/mL and the linear ranges covering two orders of magnitude with the limits of quantitation of 0.05-0.1 μg/mL. Moreover, the developed method was applied for the glycosides analysis of three tobacco samples, which only took about 2 s for every sample.

A simple method has been developed for the simultaneous determination of 16 polycyclic aromatic hydrocarbons (PAHs) in mainstream cigarette smoke. The procedure is based on employing a homemade graphene-coated solid-phase microextraction (SPME) fiber for extraction prior to GC/MS. In comparison to commercial 100-μm poly(dimethyl siloxane) (PDMS) fiber, the graphene-coated SPME fiber exhibits advantageous cleanup and preconcentration efficiencies. By collecting the particulate phase 5 cigarettes, the LODs and LOQs of 16 target PAHs were 0.02-0.07 and 0.07-0.22 ng/cigarette, respectively, and all of the linear correlation efficiencies were larger than 0.995. The validation results also indicate that the method has good repeatability (RSD between 4.2% and 9.5%) and accuracy (spiked recoveries between 80% and 110%). The developed method was applied to analyze two Kentucky reference cigarettes (1R5F and 3R4F) and six Chinese brands of cigarettes. In addition, the PAH concentrations in the particulate phase of the smoke from the 1R5F Kentucky cigarettes were in good agreement with recently reported results. Due to easy operation and good validation results, this SPME-GC/MS method may be an excellent alternative for trace analysis of PAHs in cigarette smoke.

Glycosides in tobacco leaves are highly important aromatic precursors. It is necessary to reveal glycosides in tobacco leaves to improve tobacco planting and processing. This study describes a method for the systematic screening of glycosides in tobacco leaves by liquid chromatography with tandem mass spectrometry. Although glycosides contain numerous aglycones, the number of glycans is limited. Based on a screening table of glycans designed for neutral loss scan, glycosides with different aglycones were systematically screened out. Then, the MS2 fragment spectra of scanned glycosides were further obtained using product ion scan. By comparison with the spectra in online tandem mass spectral databases, reported references, and verification by commercial standards, 64 glycosides were detected, including 39 glycosides linked with monosaccharides, 18 glycosides linked with disaccharides and 7 glycosides linked with trisaccharides. It is noteworthy that glycosides linked with trisaccharides have previously been rarely reported in tobacco. This method appears to be a useful tool for the systematic screening and characterization of glycosides in tobacco and can potentially be applied to other plants.

In this paper, heart-cutting two-dimensional GC/MS (GC-GC/MS) method in combination with a simple sample collection procedure was developed for the determination of 6 nitroalkanes in mainstream cigarette smoke. The method could remove large amounts of impurities on-line in the first polar column by heart-cuts and separate from the left interferences in a second mid-polar column. And the target compounds could be focused at the inlet of the second column by cryo-concentration. Compared to conventional GC/MS, GC-GC/MS achieved a lower noise level and sensitivity at least an order of magnitude higher. Furthermore, the GC-GC/MS method could avoid the false negative and false positive results that appeared in the compared conventional GC/MS analysis. By trapping the vapor phase of 20 cigarettes smoke, the LODs and LOQs of the nitroalkanes were 1.3 to 9.8 and 43 to 32.6 ng/cigarette, respectively, and all linear correlation efficiencies were larger than 0.999. The validation results also indicate that the method has high accuracy (spiked recoveries between 84% and 102%) and good repeatability (RSD between 7.2% and 9.4%). The developed method was applied to analyze 1 Kentucky reference cigarette (3R4F) and 10 Chinese commercial brands of cigarettes. The research results indicated that nitromethane, nitroethane, 2-nitropropane and 1-nitro-n-pentane were detected in mainstream cigarette smoke, but 1-nitro-n-butane and 2-nitropropane, which were reported by one previous study, were not detected in all cigarette samples. (C) 2015 Published by Elsevier B.V.

Urinary aromatic amines (AAs) could be used as biomarkers for human exposure to AAs in cigarette smoke. A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed for the determination of urinary AAs (i.e. 1-naphthylamine (1-NA), 2-naphthylamine (2-NA), 3-aminobiphenyl (3-ABP) and 4-aminobiphenyl (4-ABP)) in smokers and nonsmokers. A molecularly imprinted polymers (MIPs) solid phase extraction (SPE) cartridge was applied to purify urine samples and no derivatization reaction was involved. Each analytes used respective stable isotope internal standards, which could well compensate matrix effect. Lower limit of detections (LODs) for four AAs were obtained and in the range of 1.5-5ngL(-1). Recovery ranged from 87.7±4.5% to 111.3±6.4% and precision were less than 9.9%. The method was applied to analyze urine samples of 40 smokers and 10 nonsmokers. The 24h urinary excretion amounts of total AAs were higher for smokers compared with nonsmokers. What's more, 1-NA, 3-ABP and 4-ABP excretion amounts showed significant differences (p<0.05) between smokers and nonsmokers.

To elucidate the formation process of HCN from the pyrolysis of glycine, the small molecule gaseous pyrolysates, H2O, NH3, CO2, CO, HNCO, and HCN, were analyzed in real-time by TG-FTIR. The appearance of the volatile pyrolysis products and the solid residue was determined in real-time at their corresponding formation temperatures by online Py-two-dimensional GC–MS with heart-cutting and LC–MS/MS. The pyrolysis of 2,5-diketopiperazine, a thermolytic by-product of glycine pyrolysis, was also studied. The results showed that: (1) the pyrolysis of glycine can be divided into three temperature ranges 200–300, 300–440, and 440–900 °C; HCN forms in each range with three peaks appearing at 273, 422, and 763 °C, respectively. (2) The mechanistic pathways of HCN formation from glycine in the low- and high-temperature heating stages are different. Below 273 °C, glycine undergoes a decarboxylation reaction to produce methylamine, which subsequently forms HCN by means of dehydrogenation. Above 300 °C, glycine gives relatively large amounts of HCN via 2,5-diketopiperazine and subsequent HNCO or methylenimine formation.

Integration of normal-phase LC (NPLC) fractionation with RPLC-ESI/MS was established to detect the polar fractions of cuticular components from Oriental tobacco leaf. NPLC was selected for the fractionation of polar components of cuticular leaf extract, after being concentrated with rotary evaporator, each of the enriched fractions was further analyzed by RPLC-ESI/MS. In total, 83 compounds were finally detected including 45 cembranoids, 15 labdanoids, 20 sucrose esters, and 3 glucose esters (or fructose esters). Three cembranoids and seven labdanoids possibly are new diterpenoids. Glucose esters (or fructose esters) are also reported in Nicotiana tobacco for the first time.

A method based on liquid chromatography tandem mass spectrometry was developed for the direct determination of nicotine, cotinine, trans-3′-hydroxycotinine, their corresponding glucuronide conjugates as well as nornicotine, norcotinine, cotinine-N-oxide and nicotine-N′-oxide in the urine of smokers. The assay only involves centrifugation and filtration of diluted urine. The analysis was performed on a C18 reversed-phase column using a gradient of 10 mM ammonium acetate, pH 6.8, and methanol as mobile phase at a flow rate of 1 mL min−1. Nicotine-methyl-d3, Cotinine-methyl-d3 and trans-3′-hydroxycotinine-methyl-d3 were used as internal standards. Precisions (RSD) for all the analytes at three levels were between 2.1 and 17.0%. Recoveries for nicotine and nine nicotine metabolites ranged from 78.4 to 115.6%. The described method was suitable for determining the nicotine dose in large-scale human biomonitoring studies.

A rapid and accurate method for the quantification of cis-abienol in oriental tobacco leaves by normal phase liquid chromatography was developed. Freeze-dried tobacco samples were sonicated in methylene chloride for 10 min. The supernatant was purified using a silica gel solid phase extraction cartridge. Ten milliliter of the resulting methylene chloride eluate was collected, then separated on a 250 × 4.6 mm, 5 μm particle-size CN column with n-hexane: ethyl acetate, 100:2 (v/v) at a flow rate of 1 mL min−1. cis-Abienol was detected by UV absorption at 254 nm. The linear range was from 2.14 × 10−4 to 4.28 × 10−2 mg mL−1 and the correlation coefficient was 1.000. The average recovery was 98.7, 105.2 and 103.1% in five replicated sets of tobacco samples spiked with 0.2856, 0.7140 and 1.904 mg cis-abienol. The relative standard deviations (RSDs) were 1.04, 0.63 and 1.25%, respectively (n = 5). Limit of detection (S/N = 3) was 21.84 μg g−1 and limit of quantification (S/N = 10) was 72.80 μg g−1. The method was found to be suitable for determination of cis-abienol in oriental tobacco leaves. Furthermore, pure cis-abienol used for method validation was obtained by preparative reversed phase high-performance liquid chromatography. Identification was performed by UV detection, nuclear magnetic resonance and mass spectrometry.

A rapid and sensitive LC-MS/MS method was developed for the quantitative determination of sucrose esters (SEs) in Oriental tobacco samples. The sample preparation involved a 10-min sonication extraction procedure with acetone and five-fold dilution of the extract with methanol. The experiment was carried out in positive ion mode by ESI IT mass spectrometer. Because of lack of authentic standards of SEs, sucrose octa-acetate (internal standard, IS) was used as a surrogate to validate the proposed method. Matrix-matched standard calibration was used for quantification of IS in the spiked samples. Under optimized MS/MS conditions, an LOQ of 3.9 microg/g was achieved for IS, with an LOD of about 1.2 microg/g. Recoveries for IS were 95-97%. Among 19 monitored SEs, the contents of 11 SEs had RSDs lower than 13.7%. The method, with very little sample handling and good sensitivity, was applied to the rapid quantification of SEs in four Oriental tobacco samples. It appears that the sum of contents of the five SEs with MW 650, 664, and 678 Da occupied approximately 80% of the total content of SEs.

Sucrose esters (SEs) from oriental tobacco are normally characterized by gas chromatography/mass spectrometry (GC/MS) after a long saponification and derivatization procedure. To simplify the process, a rapid method has been developed by using liquid chromatography coupled with electrospray ion trap mass spectrometry (LC/ESI-MSn). Using the characteristic fragmentation behavior of abundant SEs identified by GC/MS after purification by gel permeation chromatography (GPC) from cuticular waxes of green oriental tobacco leaf, two types of SEs from green and cured oriental tobacco were identified by MSn analysis. The first is one of three types reported formerly and has 13 SE homologues. However, the presence of unsaturation in one of the acyl substituents of this first type gave rise to a new series with three homologues. The other was found to be a new type and had three homologues. The proposed method enables the rapid and sensitive characterization of SEs from oriental tobacco.

In this paper, the safe quality of Wuhe Dipsacus root as medical plant was evaluated by the analysis of the contents of zinc, lead, cadmium and mercury in Wuhe Dipsacus in the different growth years, so as to provide scientific basis for the further development and utilization of Wuhe Dipsacus Root. The results showed that those limited elements had adequate safety because their contents were much lower than the GB, and Wuhe Dipsacus were suitable for practical GAP .