Quantitative Estimation of Menthol, Glycerol and 1,2-Propylene Glycol in Tobacco Products Available in India

Quantitative Estimation of Menthol, Glycerol and 1,2-Propylene Glycol in Tobacco Products Available in India | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Article Quantitative Estimation of Menthol, Glycerol and 1,2-Propylene Glycol in Tobacco Products Available in India Prasenjit Saha, Doli Rani Das, Abhishek Kumar Yadav, Sanjib Kumar Sarma, and 2 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6161281/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Different chemicals are added to smoke and smokeless tobacco products to make the products more attractive and palatable. In this article, 43 smoke and 84 smokeless tobacco products were investigated to detect and quantify menthol, glycerol, and 1,2-propylene glycol. The data obtained were compared across the tobacco products to understand variation of these chemicals in different categories of tobacco products. The smokeless tobacco products zarda, chewing tobacco, khaini and snus contain menthol as an additive. Some smokeless tobacco such as chewing tobacco, zarda, snus and kiwam contain glycerol and / or 1,2-propylene glycol as additives. One smokeless tobacco products contain 45% glycerol which is exceptionally high compared to reported glycerol level in other studies. Among the tested 28 number of cigarette samples 39% of cigarette samples contain menthol as an additive. Most cigarettes contain both glycerol or 1,2-propylene glycol or one of them as humectants whereas bidi do not contain menthol and 1,2–propylene glycol. The determination of these additives present in the tobacco products may be helpful in further understanding the health hazards associated with these products for tobacco users. The findings in this work may provide valuable information / data to regulate the additive content in tobacco products for effective tobacco control in India. Health sciences/Medical research Physical sciences/Chemistry Figures Figure 1 Figure 2 Introduction Eight million people die on a yearly basis globally due to the consumption of any form of tobacco products. 1 Half of the tobacco users die prematurely because of tobacco related illness. Tobacco related diseases cause a huge economic burden on the world. India spends USD 27.5 billion in health care costs due to tobacco use in the year 2017-18 for persons aged 35 years and above. 2 In India 28.6% adults of age 15 and above use any form tobacco product whereas 21.4% of adults use smokeless tobacco products (SLT) and 10.7% of adults use smoke tobacco products. 1 Smoke tobacco products are consumed by inhaling the smoke from burning tobacco present in cigarettes, bidi, water pipes, cigars etc. The basic components of smoke tobacco products such as cigarettes, include tobacco, chemical additives, a filter, and paper wrapping. Approximately 8,000 different chemicals have been identified in tobacco smoke, many of which are known carcinogens and can have serious health impacts. 3 Another form of tobacco product is smokeless tobacco (SLT), which is a mixture of processed tobacco leaves with various chemicals, additives, including flavorings, areca nut, and slaked lime. 4 Smokeless tobacco products can be chewed, sucked, applied to the teeth and gums or inhaled as ground tobacco into nasal cavity. 4 – 6 Over 140 countries are affected by smokeless tobacco, raising global public health concerns, and India, with 200 million SLT users, leads both in the production and consumption of SLT products. 5 The tobacco industry are seemed to present the SLTs as a safer alternative to smoking. 7 However the use of smoke and smokeless tobacco products for prolong time can lead to nicotine addiction and cause several diseases, such as lung cancer, oral and pancreatic cancer, respiratory disease, cardiovascular diseases, chronic obstructive pulmonary disease (COPD) and various cancers. 3 – 8 Considering the enormity of the problem, member nations of the WHO negotiated and adopted the first public health treaty – the Framework Convention on Tobacco Control (FCTC) – in 2003. India was the eighth country to ratify the treaty, which provides a vital opportunity to reduce the global burden of tobacco. The Government of India launched the National Tobacco Control Programme (NTCP) in the year 2007-08 to initiate the tobacco control policies. 9 For the implementation of the tobacco control policies the three tobacco testing laboratories were set up at three different locations in India. These laboratories are involved in generating scientific data for the regulation of the content and emission of tobacco products. As per WHO guidelines on tobacco product regulation, the negative health impact of tobacco products is attributed to their toxicity, attractiveness and addictiveness. 10 Numerous compounds are added to smoke and smokeless tobacco products to increase their addictiveness and attractiveness. For example, flavors, sugars, humectants, salts and alkaline agents are used as additives in the tobacco products. 11 Since, tobacco products are inherently harmful, the addition of certain additives can pose an additional health risk to consumers. 12 Menthol is one of the most widely used flavors in both smoke and smokeless tobacco products. 11 This chemical has cooling properties that help to mask the harshness and irritation of the tobacco products. 13 – 15 The mentholated tobacco products attract the youth and young adults, often leading to smoking initiation. 14 , 15 Celebucki et al. reported that the menthol content is higher in lower-yield cigarette (nicotine content less than 6 mg/cigarette) than the regular-yield cigarette (Nicotine content 10–12 mg/cigarette). 16 Kreslake et al. noted in their study that menthol cigarettes may be adopted by first time users or occasional smokers due to their mild properties, which cover the harshness of tobacco, eventually leading to a stronger desire for the menthol flavor. 17 , 18 Brinkman and colleagues, in a cross-over study, reported the presence of higher levels of nicotine in the mouths of people who smoke menthol cigarettes, though no difference was observed in urine cotinine levels. 19 These products are also associated with increased experimentation and trial use, higher levels of dependence, and difficulty of quitting tobacco. 20 , 21 Mendez et al. in his study indicated that in the US market from 1980 to 2018 declining in smoking prevalence is only 2.6% (13.7% vs 11.1% in 2018) and estimated around 3 million deaths with 378000 premature deaths during the period 1980–2018. 22 As per the report, the low decline in smoking prevalence is due to the prevalence of mentholated cigarette which is responsible for initiation of 10.1 million new smokers. Mills et al. published a review article on the impact of menthol cigarette bans by analyzing various published database up to 2022. 14 They concluded their study by suggesting that smoking cessation rates were high after a ban on mentholated cigarette. Thus continuous monitoring and surveillance of menthol cigarettes are crucial from the perspective of public health. Glycerol and 1,2-propylene glycol are the most commonly added compounds used as humectants by the tobacco industry and often present at levels (~ 1–5% w/w) in tobacco filler of cigarettes. 23 These compounds are used for the purposes such as to retain moisture, lower the water activity and to reduce the microbial growth. 11 , 24 – 26 In smoke tobacco products these chemicals serve as the precursors of harmful carbonyl compounds such as formaldehyde, acrolein, acetaldehyde etc. 24 Literature study reveals that on burning a cigarette, almost all glycerol are pyrolysed with formation of acrolein and glycolaldehyde in trace quantity whereas approximately 15–20% of 1,2-propylene glycol are converted into 1,3-propylene glycol, acetol (hydroxyacetone), acetic anhydride, and pyruvaldehyde. 3 , 27 These chemicals are detected in both mainstream and/or side stream smoke. 28 According to our findings, smokeless tobacco products contains glycerol and 1,2-propylene glycol as humectants in different levels. Glycerol is also widely used in food products and personal care items and the toxicological studies generally indicate that glycerol is safe for these applications, especially since the majority of research focuses on oral and dermal exposure. 25 However, investigating the levels of such humectants in SLTs and their harmful effects is of immediate concern. Recently, the WHO Study Group on Tobacco Product Regulation (TobReg.) has indeed highlighted the importance of regulating toxicants in tobacco products and proposed mandated lowering of different toxicants viz. formaldehyde, acetaldehyde, acrolein, 1,3-butadiene, etc. 3 , 24 Pennings et al. carried out the correlation study of the amounts of aldehydes, VOCs, and CO present in mainstream smoke of 50 commercially available cigarette brands with their potential precursor’s sugars and humectants present in the cigarette filler. 3 They stated that reduction of emission of chemicals such as formaldehyde, acetaldehyde, acrolein and 2,5-dimethyfuran from cigarette smoke may occurs when sugar and humectants contents are lowered in tobacco filler. Gupta et al. studied the level of sugars, humectants, and their toxic by-products in various tobacco products. 24 The authors mentioned that smokeless tobacco products sold in South Asia such as zarda, chewing tobacco, gutkha, and khaini contain high levels of sugars and humectants. So systematic study of tobacco additives like humectants or chemicals formed during their combustion should be prior concern as users of such unregulated additive present in tobacco products are vulnerable to potentially harmful health effects. In India, different varieties of smokeless tobacco products are available for human consumption. Among them chewing tobacco, zarda, sada, khaini, snus, sada, gul, gurakhu, snuff, kiwam etc. are the most commonly used smokeless tobacco products in India. 29 , 30 Cigarettes and bidi are more popular smoke tobacco products sold and consumed in India. Herein, we investigated and reported the quantitative estimation of menthol, glycerol and 1,2-propylene glycol in smoke and smokeless tobacco products which contribute attractiveness and toxicity of the tobacco products. Methods The tobacco samples used for analysis were received from 12 states in India, from the different District or State Tobacco control cells under the National Tobacco Control Program (NTCP), Government of India. The sample selection was done randomly as the laboratory received from state nodal officers (SNO), NTCP. SNO collect the samples from the market so that the most consumed and prevalent tobacco products are selected by them for the laboratory analysis. In this investigation 84 number of smokeless tobacco samples, 28 number of cigarette samples and 15 number of bidi samples were used. After receiving the tobacco samples, the samples were stored at -20°C. Before the analysis the samples were kept at refrigerator for minimum 24 hours and then the samples were kept at room temperature for 1 h. For the determination of menthol and humectants in smoke and smokeless tobacco products, CORESTA recommended method No. 92 and WHO TobLabNet SOP 06 were followed respectively (The detail procedures followed are described in supplementary information and these methods are also available online). 31 , 32 CP wax 51 or DB wax and DB wax or BP-624 capillary GC column were used respectively for the estimation of menthol and humectants in the samples using Gas Chromatograph with FID detector,. 31 , 32 The laboratory is accredited by NABL as per ISO/IEC 17025: 2017 for the testing of menthol, glycerol and 1,2-propylene glycol. Results Menthol The smokeless tobacco products chewing tobacco, zarda, khaini, pouch khaini, sada, gul, gurakhu samples were analyzed for the presence and quantification of menthol flavor. The menthol content in 35 brands of chewing tobacco/ zarda (smokeless) were found in the range of 1.6–27.0 mg/g tobacco. (Table 1 , Sl. No. 1; See Suppl. Information for further details). However, menthol could not be traced in smokeless tobacco products such as gul (number of samples, n = 10), sada (n = 3), gudakhu (n = 7), snuff (n = 3) and khaini (n = 6) products (Table 2 , Suppl. Information). Moreover, 3 brands of khaini and sada samples contain trace level (0.23–0.34 mg/g) of menthol in it. 8 brands of khaini & sada products such as Raja Tobacco, Afgan Rally, Chunnu Right Choice, Golden Sada etc. and 2 brands of snus products such as Chaini khaini, Bolt Filter contain menthol as an additive (Fig. 1 ). The amount of menthol in these 8 brands of khaini & sada products were present in the range of 1.5–8.7 mg/g tobacco (Fig. 1 ). Two kiwam products were tested and menthol was present in both the samples in the range of 13.0–24.9 mg/g tobacco. The laboratory has quantified the menthol content in various cigarette and bidi samples (Tables 3 and 4, Suppl. Information). Some cigarette brands such as Gold Flake Indie Mint, Wave Cool Mint, Total, Wave Fruit Mint, Topaz, Players contain menthol ball/capsule 33,34 inside the cigarette filter which are broken by pressing the filter of these types of cigarettes during smoking by the consumer. Menthol was estimated in cigarette as mg/cigarette as two whole cigarette were taken for analysis as per the CORESTA standard method. The laboratory analyzed 28 numbers of different brands of cigarettes and among them 11 brands of cigarette contain menthol in the range of 0.02–9.3 mg per cigarette (Fig. 2). It indicates that around 39% of cigarette samples sold in Indian market contain menthol as per the present study is concerned. Our laboratory has tested 15 number of different brands of bidi samples and menthol is found in the range of ND – 0.21 mg/g of tobacco filler of bidi sample (Table 1 ; Sl. No. 11). Table 1 Menthol content in different brands of smokeless tobacco samples and smoke tobacco samples: Sl. No. Type of Tobacco Sample Smokeless / Smoke Tobacco No. of Brands analyzed Menthol Unit 1 Chewing Tobacco & Zarda Smokeless 35 1.6–27.0 mg/g 2 Snus Smokeless 2 7.6–8.3 mg/g 3 Khaini Smokeless 14 ND – 8.7 mg/g 4 Sada Smokeless 6 ND – 3.5 mg/g 5 Gul Smokeless 10 ND mg/g 6 # Dried leaf Smokeless 5 ND mg/g 7 Gurakhu Smokeless 7 ND mg/g 8 Snuff Smokeless 3 ND mg/g 9 Kiwam Smokeless 2 13.0–24.9 mg/g 10 Cigarettes Smoke 28 ND – 9.3 mg/cig. 11 Bidi Smoke 15 ND – 0.21 mg/g ND: Not Detected. (Limit of Detection of Menthol Glycerol = 0.013 mg/g). # Received from different sources. Humectant (Glycerol and 1,2-Propylene Glycol) The most commonly used humectants in tobacco products are glycerol and 1,2-propylene glycol. 23 – 26 This laboratory has tested a variety of smokeless tobacco products and among them zarda, chewing tobacco and kiwam contain high levels of glycerol and 1,2 – propylene glycol. Mean glycerol and 1,2- propylene glycol were found in the range of 0.9–185.3 mg/g and ND – 40.8 mg/g respectively (Table 2 ; See Table 5 Suppl. Information for more details)). A kiwam samples Raj Ratan tobacco contain exceptionally high quantity of glycerol (456.28 mg/g tobacco) whereas Royal Tulsi contain both glycerol (2.3 mg/g tobacco) and 1,2-propylene glycol (23.9 mg/g tobacco) (Table 6, Suppl. Information). 10 number of gul samples were analysed for the presence and quantification of humectants (Table 6, Suppl. Information). Among them glycerol was present in 1 gul samples (1.12 mg/g) whereas another 1 gul sample contain 1.11 mg/g of 1,2-propylene glycol. Other smokeless tobacco products such as snuff, gurakhu, dried leaf and sada do not contain any glycerol and 1,2-propylene glycol (Table 6, Suppl. Information). Among 14 khaini samples tested in this laboratory, 7 samples do not contain glycerol, 4 samples contain trace quantity of glycerol which is below the limit of quantitation and 3 samples contain glycerol in the range of 1.23–4.56 mg/g tobacco (Table 6, Suppl. Information). However among 14 khaini samples 1,2 – propylene glycol was absent in 13 khaini samples and 1 khaini sample contain 0.64 mg/g tobacco. Table 2 Determination of Additives (Glycerol and 1,2-Propylene Glycol) in different variety of smokeless and smoke tobacco samples. Sl. No. Tobacco Sample Types of Tobacco Sample No. of Brands analyzed Glycerol (mg/g) PG (mg/g) 1 Chewing Tobacco & Zarda SLT 35 0.9–185.3 ND – 40.8 2 Snus SLT 2 39.5–53.4 2.1–16.6 3 Gul SLT 10 ND – 1.12 ND − 1.11 4 Khaini SLT 14 ND – 4.56 ND – 0.64 5 Sada SLT 6 ND ND 6 # Dried leaf SLT 5 ND ND 7 Gurakhu SLT 3 ND ND 8 Snuff SLT 3 ND ND 9 Kiwam SLT 2 2.26–456.3 ND – 23.9 10 Cigarette Smoke 28 ND − 22.3 ND – 10.7 11 Bidi Smoke 15 ND − 3.6 ND ND: Not Detected. BLOQ: below limit of quantitation. (Limit of Quantitation of Glycerol = 0.9 mg/g and 1,2-Propylene Glycol = 0.2 mg/g). # Received from different sources. Among smoke tobacco samples, bidi and cigarettes samples were analyzed for the quantification of glycerol and 1,2-propylene glycol (PG). Out of 28 cigarette samples tested, 27 number of cigarette samples contain glycerol or 1,2-propylene glycol as one of the additives. The amount of glycerol and 1,2-propylene glycol was present in cigarette sample ranges from not detected (ND) to 22.33 mg/g tobacco (Mean = 10.2 mg/g) and ND to 10.70 mg/g tobacco (Mean = 2.2 mg/g) respectively (Table 7, Suppl. Information). Among the cigarette samples tested, only 4 cigarette samples P-10 premium, Charms, H-10 and Sunils Filter did not contain glycerol as an additive whereas 3 samples did not contain 1,2-propylene glycol and 8 samples contain trace quantity of 1,2-propylene glycol which is below limit of quantitation. Among the 15 bidi brands tested in our laboratory, 1,2-propylene glycol was not detected in all samples whereas glycerol was not present in 7 samples. 8 bidi samples contain glycerol in the range of 1.1–3.6 mg/g (Mean = 1.9 mg/g tobacco). Discussion Smokeless tobacco products sold in India contain high levels of flavors and sweetening agent to enhance the palatability of tobacco product. Among the flavors menthol is most extensively used chemicals in tobacco products due to their cooling effect which help to mask the harshness of the products. 20 , 35 , 36 Lisko et al. in 2014 studied the levels of flavors in smokeless tobacco products of South East Asia of varieties like zarda, quiwam, gutkha, and khaini samples where menthol was present in zarda samples in the range 5–22 mg/g tobacco. 12 Cheah and coworkers recently carried out the study of analysis of composition of smokeless tobacco products (zarda/chewing tobacco, khaini, kharra and gudakhu) sold in north, east west and central regions of India. Menthol was measured in two zarda samples and found in the range of 4.1–9.7 mg/g tobacco whereas in three chewing tobacco, menthol was found in the range of 0.3–2.0 mg/g tobacco. 4 In the present study, menthol was detected in all the zarda and chewing tobacco samples and was found in the range of 1.6–27.0 mg/g tobacco, which is consistent with the findings of Lisko & coworkers. 12 In 2018, Stanfill et al. investigated the composition of packaged SLT products available in a well-established South Asian community outside of South Asia. In their work menthol content was reported in the range of 0.16–21.7 mg/g tobacco and 7.0–11.0 mg/g tobacco respectively for zarda and khaini samples which are comparable to the present study where samples are mostly collected from North-Eastern part of India. 30 From the observations, it can be inferred that smokeless tobacco products sold in Indian market contains varying menthol concentrations; upto 17 fold in zarda & chewing tobacco from lowest quantity quantified in zarda & chewing tobacco sample. In our investigation, the snus products Chaini khaini, Bolt Filter contain menthol in the range of 7.6–8.3 mg/g tobacco. Stepanov et al. mentioned in their brief report that Chaini khaini is marketed as a safe alternative to other tobacco products despite of containing high levels of carcinogenic nitrosamines and biologically available nicotine. 37 Stanfill and coworkers investigated the menthol content in four kiwam products and contain menthol (0.85–12.3 mg/g tobacco) in all products. 30 In our present study, two samples were investigated which contain menthol (13.0-24.9 mg/g tobacco). The consumption of such mentholated flavored tobacco may enhance the addiction towards such harmful products and regulating or banning menthol additives should have immediate focus for better public health. In the present study, the menthol content in mentholated cigarettes was found in the range of 2.3–9.3 mg/cigarette (Table 3, Suppl. Information). Taylor et al . has studied the menthol content in US marketed cigarettes and found that menthol content in 23 menthol branded cigarettes in the range of 2.9–19.6 mg/cigarette whereas in non-mentholated cigarette menthol was present in the range of 0.002–0.07 mg/cigarette. 38 The observations of our laboratory were consistent with the findings of Taylor & coworkers but highest menthol content in mentholated Indian cigarette is 9.3 mg/cigarette which is somewhat lesser than US marketed cigarettes which is 19.6 mg/cigarette. Silva et al. reported in their article that tobacco companies use menthol in 'non-menthol' cigarettes at subliminal levels, i.e., at levels not perceptible as menthol-flavored. 21 In this investigation, in two non-mentholated cigarettes viz. silk cut and gold flake premium, menthol was found in the range of 0.02–0.07 mg/cigarette (Fig. 2) which are at subliminal level. Wu group mentioned that bidi cigarettes sold in the USA contain various flavorings, including vanilla, chocolate, strawberry, grape, root beer and others whereas Indian bidis do not contain any flavor additives. 39 , 40 Stanfill & coworkers reported that menthol was present in the range of ND – 2.77 mg/bidi in 10 of the flavored bidi brands including non-mentholated brands. 40 Our laboratory has tested 15 number of different brands of bidi samples where menthol could not be traced in 11 samples (Table 4, Suppl. Information). In two bidi samples menthol was present in the range of 0.15–0.21 mg/g tobacco whereas in another two samples menthol was detected which was below limit of quantification. (LOQ = 0.04 mg/g). From this studies, it can be inferred that most Indian bidis do not contain menthol additives. The additives glycerol and 1,2-propylene glycol, used as humectants in tobacco products, contain varying levels of these substances. Gupta et al . in their report mentioned that the chewing tobacco products contain high levels of humectants, with mean glycerol levels between 24 to 40 mg/g tobacco and propylene glycol between 1.9 to 8.0 mg/g tobacco. 24 In our findings seven chewing tobacco products and zarda contain exceptionally high quantities of glycerol around 2–5 fold higher than the high value of humectants found in most of the chewing tobacco products (Table 5, Sl. No. 29–35, Supplementary Information). Foulds et al. and Rickert et al. reported that Swedish snus contains 15–35 mg/g humectants (e.g. propylene glycol and glycerol) and 16.2–16.4 mg/g 1,2-propylene glycol resepectively. 41 , 42 In our investigation snus products such as Chaini filter pouch, Bolt Filter contain glycerol and 1,2 – propylene glycol in the range of 39.5–53.4 mg/g tobacco and 2.1–16.6 tobacco mg/g respectively (Table 6, Supplementary Information). Rickert et al . studied the presence of glycerol and 1,2-propylene glycol in 25 brands of snuff products and found that glycerol was absent in all products whereas 1,2-propylene glycol was present in only 7 samples in the range of 3.92–23.4 mg/g tobacco. 42 However, in our investigation all the three snuff products do not contain glycerol and 1,2-propylene glycol. In our findings the amount of glycerol and 1,2-propylene glycol in cigarette samples were present in the range of ND – 22.33 mg/g tobacco and ND – 10.70 mg/g tobacco. Pennings and coworkers measured the quantity of glycerol and 1,2-propylene glycol in 52 tobacco samples and found that these substances were present in the range of 1.1–23.7 mg/g of tobacco (mean = 7.8 mg/g) and 0–14.8 mg/g of tobacco (mean = 4.6 mg/g), respectively. 3 In our investigation, the maximum amount of 1,2-propylene glycol in cigarettes was found to be 10.7 mg/g tobacco which is lower than the findings of the Pennings group, i.e. 14.8 mg/g tobacco. It was also observed that glycerol was used as a humectant in most cigarettes more frequently than 1,2-propylene glycol. The amount of glycerol (1.06–22.33 mg/g tobacco, mean = 10.2 mg/g) added in cigarettes are also comparatively higher than 1,2-propylene glycol (0.3–10.7 mg/g tobacco; mean = 2.2 mg/g). Most cigarettes contain glycerol and/or 1,2-propylene glycol as additives, which makes the product more toxic, as these additives produce harmful aldehydes when burned. In comparison of cigarette and bidi products, cigarette products contain high quantity of glycerol almost 7 fold higher compared to bidi products (Table 8, Suppl. Information). Chewing tobacco/ zarda and kiwam products contain exceptionally high quantity of glycerol upto 18% and 45% respectively compared to the cigarettes which contain upto 2.2% glycerol. Conclusion According to recommendations from the WHO TobReg, it is crucial to implement product standards and strategies to mandate the reduction of toxicants in all types of tobacco products by removing or lowering the contents such as menthol and humectant. 24 , 43 The contents menthol, glycerol and 1,2-propylene glycol in smoke and smokeless tobacco products available in India are poorly studied and so far received limited attention from researchers and policy makers. The above findings may be useful to regulate the additives in the tobacco products to minimize the exposures and risks associated with the use of smoke and smokeless tobacco products. Declarations Competing Interests The authors declare no conflict of interests and no financial interests. Author Contribution The experimental work was carried out by D.R.D., A.K.Y., S.K.S. and K.J.B. at NTTL Guwahati. The results were analyzed by P.S. The main manuscript text was prepared by P.S. and D.R.D. The manuscript text was reviewed and edited by D.K. The manuscript was reviewed by all authors Acknowledgement The finding and conclusions reported in this article are those of the author(s) and do not necessarily represent the views of the National Tobacco Control Program (NTCP), Ministry of health & Family Welfare, Government of India. Brand names are used for identification only and the laboratory does not endorse any tobacco product. We greatly acknowledge Dr. L. Swasticharan, Addl. DDG, NTCP, State Nodal Officers, NTCP and National Tobacco Control Program, Ministry of Health & Family Welfare, Government of India for the support in this investigation. Data Availability The data generated are incorporated into the article and its online supplementary material. References Tata Institute of Social Sciences (TISS), Mumbai and Ministry of Health and Family Welfare, Government of India. 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Smokeless tobacco and cigarette smoking: chemical mechanisms and cancer prevention. Nat. Rev. Cancer 22 (3), 143–155 (2022). Chen, X., Shi, Q., Zhou, X. & Liu, X. Molecular characterization of aldehydes and ketones in particle phase of mainstream and sidestream cigarette smoke. R Soc. Open Sci. 6(2), 181832 (2019). Smokeless tobacco (SLT) products - 2018. WHO FCTC Knowledge Hub. https://extranet.who.int/fctcapps/fctcapps/fctc/kh/slt/news/smokeless-tobacco-slt-products. Stanfil,l S. B. et al . Chemical characterization of smokeless tobacco products from south asia: nicotine, unprotonated nicotine, tobacco-specific n-nitrosamines, and flavor compounds, Food Chem. Toxicol. 118 , 626-634 (2018). Determination of menthol in cigarettes and cut filler by gas chromatography. CORESTA Recommended Method No. 92 (2019). Standard operating procedure for the determination of humectants in cigarette tobacco filler. WHO TobLabNet Official Method SOP 06; (2016). Mus S, et al. , Chemicals in cigarette flavor capsules from guatemala and mexico. Nicotine Tob. Res. 26 (5), 545-551 (2024). Kim, H. S. et al. Quantitative analysis of menthol and identification of other flavoring ingredients in capsule cigarettes marketed in Korea. Regul Toxicol Pharmacol . 92 , 420-428 (2018). Wayne, G. F. & Connolly, G. Application, function and effects of menthol in cigarettes, a survey of tobacco industry documents. Nicotine Tob Res . 6 , 43-54 (2004). Chen, C., Isabelle, L. M., Pickworth, W. B. & Pankow, J. F. Levels of mint and wintergreen flavorants: Smokeless tobacco products vs. confectionery products. Food Chem. Toxicol . 48 (2), 755–763 (2010). Stepanov, I. et al . High levels of tobacco-specific n-nitrosamines and nicotine in chaini khaini, a product marketed as snus. Tob. Control 24 (e4), e271-e274 (2015). Ai, J., Taylor, K. M., Lisko, J. G., Tran, H., Watson, C. H., Matthew, R. & Holman, M. R. Menthol content in US marketed cigarettes. Nicotine Tob. Res. 18 (7), 1575-1580 (2016). Wu, W., Song, S., Ashley, D. L. & Watson, C. H. Assessment of tobacco-specific nitrosamines in the tobacco and mainstream smoke of Bidi cigarettes. Carcinogenesis 25 (2), 283-287 (2004). Stanfill, S, B., Brown, C. R., Yan, X., Watson, C. H. & Ahley, D. L. Quantification of flavor-related compounds in the unburned contents of bidi and clove cigarettes. J. Agric. Food Chem . 54 (22), 8580-8588 (2006). Foulds, J., Ramstrom. L., Burke. M. & Fagerström. K. Effect of smokeless tobacco (snus) on smoking and public health in Sweden. Tob. Control 12 (4), 349–359, (2003). Rickert, W. S., Joza, P. J., Trivedi, A. H., Momin, R. A., Wagstaff, W. G. & Lauterbach, J. H. Chemical and toxicological characterization of commercial smokeless tobacco products available on the Canadian market. Regul. Toxicol. Pharmacol. 53 , 121–133 (2009). Eldridge, A.C., McAdam, K. G., Betson, T.R., Gama, M. V. & Proctor, C. J. Impact assessment of WHO TobReg proposals for mandated lowering of selected mainstream cigarette smoke toxicants. Regul. Toxicol. Pharmacol. 86 , 332-348 (2017). Additional Declarations No competing interests reported. Supplementary Files SupplementaryInformation.pdf Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. Our growing team is made up of researchers and industry professionals working together to solve the most critical problems facing scientific publishing. Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-6161281","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":443396712,"identity":"4639af67-cc23-42cf-8c99-152c6bafa0d6","order_by":0,"name":"Prasenjit Saha","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA8klEQVRIiWNgGAWjYDADAwbmAwc+ABls7MRrYUt8OAOkhZl4LTzGxjwgFiEt5tKHH3/4uKNO3pz9gJm0za9t8nzMDIwfPubg1mLZl2YmOfPMYcOdPQlp0rl9tw3bmBmYJWduw+OeMwxmzLxtBxg3HEg4Jp3bc5sRqIWNmRevFvbPn/+21dlvOP+wTdqy57Y9EVp4DKSBJiduuJHMbMzw43YiQS2WPTxlkr1th5M33HjG+LC34XZyGzNjM16/mPOwb/7ws63OdsP5/A8Hfvy5bTu/vfngh4/4HIbCY2wDkw241WNoYfiDV/EoGAWjYBSMUAAA0y1T4VsjEMoAAAAASUVORK5CYII=","orcid":"","institution":"National Tobacco Testing Laboratory Guwahati","correspondingAuthor":true,"prefix":"","firstName":"Prasenjit","middleName":"","lastName":"Saha","suffix":""},{"id":443396713,"identity":"388bec57-851a-49d6-872d-75f7dc90683a","order_by":1,"name":"Doli Rani Das","email":"","orcid":"","institution":"National Tobacco Testing Laboratory Guwahati","correspondingAuthor":false,"prefix":"","firstName":"Doli","middleName":"Rani","lastName":"Das","suffix":""},{"id":443396714,"identity":"2e6fe9fc-048e-4bd4-8aee-916b04285daf","order_by":2,"name":"Abhishek Kumar Yadav","email":"","orcid":"","institution":"National Tobacco Testing Laboratory Guwahati","correspondingAuthor":false,"prefix":"","firstName":"Abhishek","middleName":"Kumar","lastName":"Yadav","suffix":""},{"id":443396715,"identity":"27164f97-ffeb-4380-add2-5890688e92a3","order_by":3,"name":"Sanjib Kumar Sarma","email":"","orcid":"","institution":"National Tobacco Testing Laboratory Guwahati","correspondingAuthor":false,"prefix":"","firstName":"Sanjib","middleName":"Kumar","lastName":"Sarma","suffix":""},{"id":443396716,"identity":"807a677b-3e44-42f5-93e3-8ea1be8d11b1","order_by":4,"name":"Kalyan Jyoti Borah","email":"","orcid":"","institution":"National Tobacco Testing Laboratory Guwahati","correspondingAuthor":false,"prefix":"","firstName":"Kalyan","middleName":"Jyoti","lastName":"Borah","suffix":""},{"id":443396717,"identity":"44fc0c4e-a8be-4f5c-8887-4d00a7a778ef","order_by":5,"name":"Dinesh Kumar","email":"","orcid":"","institution":"National Tobacco Testing Laboratory Guwahati","correspondingAuthor":false,"prefix":"","firstName":"Dinesh","middleName":"","lastName":"Kumar","suffix":""}],"badges":[],"createdAt":"2025-03-05 09:53:12","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-6161281/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-6161281/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":81035644,"identity":"4512e6df-fe36-40f1-86d8-13a77196388f","added_by":"auto","created_at":"2025-04-21 12:09:15","extension":"jpg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":39204,"visible":true,"origin":"","legend":"\u003cp\u003eMenthol content in khaini and pouch khaini/snus products.\u003c/p\u003e","description":"","filename":"1.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6161281/v1/500339ba2124dd14a4579069.jpg"},{"id":81035646,"identity":"65682103-4b0e-44b4-ac02-962fe5be811b","added_by":"auto","created_at":"2025-04-21 12:09:15","extension":"jpg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":34171,"visible":true,"origin":"","legend":"\u003cp\u003eMenthol content in cigarettes tested in the laboratory.\u003c/p\u003e","description":"","filename":"2.jpg","url":"https://assets-eu.researchsquare.com/files/rs-6161281/v1/009c2e2d0b40c4e9cacbab74.jpg"},{"id":102827674,"identity":"b20ad3a0-4d04-4c48-977e-86ee1a84426f","added_by":"auto","created_at":"2026-02-17 09:12:04","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":724863,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-6161281/v1/3b1c3ff8-d851-413c-b347-446bcb5fb983.pdf"},{"id":81035649,"identity":"afca76d1-d155-4eba-9763-109b886d86e8","added_by":"auto","created_at":"2025-04-21 12:09:15","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"supplement","size":529452,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryInformation.pdf","url":"https://assets-eu.researchsquare.com/files/rs-6161281/v1/35fe7db5e79347acedf34593.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"Quantitative Estimation of Menthol, Glycerol and 1,2-Propylene Glycol in Tobacco Products Available in India","fulltext":[{"header":"Introduction","content":"\u003cp\u003eEight million people die on a yearly basis globally due to the consumption of any form of tobacco products.\u003csup\u003e\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u003c/sup\u003e Half of the tobacco users die prematurely because of tobacco related illness. Tobacco related diseases cause a huge economic burden on the world. India spends USD 27.5\u0026nbsp;billion in health care costs due to tobacco use in the year 2017-18 for persons aged 35 years and above.\u003csup\u003e\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e\u003c/sup\u003e In India 28.6% adults of age 15 and above use any form tobacco product whereas 21.4% of adults use smokeless tobacco products (SLT) and 10.7% of adults use smoke tobacco products.\u003csup\u003e\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003cp\u003eSmoke tobacco products are consumed by inhaling the smoke from burning tobacco present in cigarettes, bidi, water pipes, cigars etc. The basic components of smoke tobacco products such as cigarettes, include tobacco, chemical additives, a filter, and paper wrapping. Approximately 8,000 different chemicals have been identified in tobacco smoke, many of which are known carcinogens and can have serious health impacts.\u003csup\u003e\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003cp\u003eAnother form of tobacco product is smokeless tobacco (SLT), which is a mixture of processed tobacco leaves with various chemicals, additives, including flavorings, areca nut, and slaked lime.\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e\u003c/sup\u003e Smokeless tobacco products can be chewed, sucked, applied to the teeth and gums or inhaled as ground tobacco into nasal cavity.\u003csup\u003e\u003cspan additionalcitationids=\"CR5\" citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e\u003c/sup\u003e Over 140 countries are affected by smokeless tobacco, raising global public health concerns, and India, with 200\u0026nbsp;million SLT users, leads both in the production and consumption of SLT products.\u003csup\u003e\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e\u003c/sup\u003e The tobacco industry are seemed to present the SLTs as a safer alternative to smoking.\u003csup\u003e\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e\u003c/sup\u003e However the use of smoke and smokeless tobacco products for prolong time can lead to nicotine addiction and cause several diseases, such as lung cancer, oral and pancreatic cancer, respiratory disease, cardiovascular diseases, chronic obstructive pulmonary disease (COPD) and various cancers.\u003csup\u003e\u003cspan additionalcitationids=\"CR4 CR5 CR6 CR7\" citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003cp\u003eConsidering the enormity of the problem, member nations of the WHO negotiated and adopted the first public health treaty \u0026ndash; the Framework Convention on Tobacco Control (FCTC) \u0026ndash; in 2003. India was the eighth country to ratify the treaty, which provides a vital opportunity to reduce the global burden of tobacco. The Government of India launched the National Tobacco Control Programme (NTCP) in the year 2007-08 to initiate the tobacco control policies.\u003csup\u003e\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e\u003c/sup\u003e For the implementation of the tobacco control policies the three tobacco testing laboratories were set up at three different locations in India. These laboratories are involved in generating scientific data for the regulation of the content and emission of tobacco products.\u003c/p\u003e \u003cp\u003eAs per WHO guidelines on tobacco product regulation, the negative health impact of tobacco products is attributed to their toxicity, attractiveness and addictiveness.\u003csup\u003e\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e\u003c/sup\u003e Numerous compounds are added to smoke and smokeless tobacco products to increase their addictiveness and attractiveness. For example, flavors, sugars, humectants, salts and alkaline agents are used as additives in the tobacco products.\u003csup\u003e\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e\u003c/sup\u003e Since, tobacco products are inherently harmful, the addition of certain additives can pose an additional health risk to consumers.\u003csup\u003e\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003cp\u003eMenthol is one of the most widely used flavors in both smoke and smokeless tobacco products.\u003csup\u003e\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e\u003c/sup\u003e This chemical has cooling properties that help to mask the harshness and irritation of the tobacco products.\u003csup\u003e\u003cspan additionalcitationids=\"CR14\" citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e\u003c/sup\u003e The mentholated tobacco products attract the youth and young adults, often leading to smoking initiation.\u003csup\u003e\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e,\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e\u003c/sup\u003e Celebucki \u003cem\u003eet al.\u003c/em\u003e reported that the menthol content is higher in lower-yield cigarette (nicotine content less than 6 mg/cigarette) than the regular-yield cigarette (Nicotine content 10\u0026ndash;12 mg/cigarette).\u003csup\u003e16\u003c/sup\u003e Kreslake \u003cem\u003eet al.\u003c/em\u003e noted in their study that menthol cigarettes may be adopted by first time users or occasional smokers due to their mild properties, which cover the harshness of tobacco, eventually leading to a stronger desire for the menthol flavor.\u003csup\u003e\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e,\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e\u003c/sup\u003e Brinkman and colleagues, in a cross-over study, reported the presence of higher levels of nicotine in the mouths of people who smoke menthol cigarettes, though no difference was observed in urine cotinine levels.\u003csup\u003e\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e\u003c/sup\u003e These products are also associated with increased experimentation and trial use, higher levels of dependence, and difficulty of quitting tobacco.\u003csup\u003e\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e,\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003cp\u003eMendez \u003cem\u003eet al.\u003c/em\u003e in his study indicated that in the US market from 1980 to 2018 declining in smoking prevalence is only 2.6% (13.7% vs 11.1% in 2018) and estimated around 3\u0026nbsp;million deaths with 378000 premature deaths during the period 1980\u0026ndash;2018.\u003csup\u003e\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e\u003c/sup\u003e As per the report, the low decline in smoking prevalence is due to the prevalence of mentholated cigarette which is responsible for initiation of 10.1\u0026nbsp;million new smokers. Mills \u003cem\u003eet al.\u003c/em\u003e published a review article on the impact of menthol cigarette bans by analyzing various published database up to 2022.\u003csup\u003e\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e\u003c/sup\u003e They concluded their study by suggesting that smoking cessation rates were high after a ban on mentholated cigarette. Thus continuous monitoring and surveillance of menthol cigarettes are crucial from the perspective of public health.\u003c/p\u003e \u003cp\u003eGlycerol and 1,2-propylene glycol are the most commonly added compounds used as humectants by the tobacco industry and often present at levels (~\u0026thinsp;1\u0026ndash;5% w/w) in tobacco filler of cigarettes.\u003csup\u003e\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e\u003c/sup\u003e These compounds are used for the purposes such as to retain moisture, lower the water activity and to reduce the microbial growth.\u003csup\u003e\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e,\u003cspan additionalcitationids=\"CR25\" citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e\u003c/sup\u003e In smoke tobacco products these chemicals serve as the precursors of harmful carbonyl compounds such as formaldehyde, acrolein, acetaldehyde etc.\u003csup\u003e\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e\u003c/sup\u003e Literature study reveals that on burning a cigarette, almost all glycerol are pyrolysed with formation of acrolein and glycolaldehyde in trace quantity whereas approximately 15\u0026ndash;20% of 1,2-propylene glycol are converted into 1,3-propylene glycol, acetol (hydroxyacetone), acetic anhydride, and pyruvaldehyde.\u003csup\u003e\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e,\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e\u003c/sup\u003e These chemicals are detected in both mainstream and/or side stream smoke.\u003csup\u003e\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e\u003c/sup\u003e According to our findings, smokeless tobacco products contains glycerol and 1,2-propylene glycol as humectants in different levels. Glycerol is also widely used in food products and personal care items and the toxicological studies generally indicate that glycerol is safe for these applications, especially since the majority of research focuses on oral and dermal exposure.\u003csup\u003e\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e\u003c/sup\u003e However, investigating the levels of such humectants in SLTs and their harmful effects is of immediate concern. Recently, the WHO Study Group on Tobacco Product Regulation (TobReg.) has indeed highlighted the importance of regulating toxicants in tobacco products and proposed mandated lowering of different toxicants viz. formaldehyde, acetaldehyde, acrolein, 1,3-butadiene, etc.\u003csup\u003e\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e, \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003cp\u003ePennings \u003cem\u003eet al.\u003c/em\u003e carried out the correlation study of the amounts of aldehydes, VOCs, and CO present in mainstream smoke of 50 commercially available cigarette brands with their potential precursor\u0026rsquo;s sugars and humectants present in the cigarette filler.\u003csup\u003e\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e\u003c/sup\u003e They stated that reduction of emission of chemicals such as formaldehyde, acetaldehyde, acrolein and 2,5-dimethyfuran from cigarette smoke may occurs when sugar and humectants contents are lowered in tobacco filler. Gupta \u003cem\u003eet al.\u003c/em\u003e studied the level of sugars, humectants, and their toxic by-products in various tobacco products.\u003csup\u003e\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e\u003c/sup\u003e The authors mentioned that smokeless tobacco products sold in South Asia such as zarda, chewing tobacco, gutkha, and khaini contain high levels of sugars and humectants. So systematic study of tobacco additives like humectants or chemicals formed during their combustion should be prior concern as users of such unregulated additive present in tobacco products are vulnerable to potentially harmful health effects.\u003c/p\u003e \u003cp\u003eIn India, different varieties of smokeless tobacco products are available for human consumption. Among them chewing tobacco, zarda, sada, khaini, snus, sada, gul, gurakhu, snuff, kiwam etc. are the most commonly used smokeless tobacco products in India.\u003csup\u003e\u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e,\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e\u003c/sup\u003e Cigarettes and bidi are more popular smoke tobacco products sold and consumed in India. Herein, we investigated and reported the quantitative estimation of menthol, glycerol and 1,2-propylene glycol in smoke and smokeless tobacco products which contribute attractiveness and toxicity of the tobacco products.\u003c/p\u003e"},{"header":"Methods","content":"\u003cp\u003eThe tobacco samples used for analysis were received from 12 states in India, from the different District or State Tobacco control cells under the National Tobacco Control Program (NTCP), Government of India. The sample selection was done randomly as the laboratory received from state nodal officers (SNO), NTCP. SNO collect the samples from the market so that the most consumed and prevalent tobacco products are selected by them for the laboratory analysis. In this investigation 84 number of smokeless tobacco samples, 28 number of cigarette samples and 15 number of bidi samples were used. After receiving the tobacco samples, the samples were stored at -20\u0026deg;C. Before the analysis the samples were kept at refrigerator for minimum 24 hours and then the samples were kept at room temperature for 1 h. For the determination of menthol and humectants in smoke and smokeless tobacco products, CORESTA recommended method No. 92 and WHO TobLabNet SOP 06 were followed respectively (The detail procedures followed are described in supplementary information and these methods are also available online).\u003csup\u003e\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e,\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e\u003c/sup\u003e CP wax 51 or DB wax and DB wax or BP-624 capillary GC column were used respectively for the estimation of menthol and humectants in the samples using Gas Chromatograph with FID detector,.\u003csup\u003e\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e,\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e\u003c/sup\u003e The laboratory is accredited by NABL as per ISO/IEC 17025: 2017 for the testing of menthol, glycerol and 1,2-propylene glycol.\u003c/p\u003e"},{"header":"Results","content":"\u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003eMenthol\u003c/h2\u003e \u003cp\u003eThe smokeless tobacco products chewing tobacco, zarda, khaini, pouch khaini, sada, gul, gurakhu samples were analyzed for the presence and quantification of menthol flavor. The menthol content in 35 brands of chewing tobacco/ zarda (smokeless) were found in the range of 1.6\u0026ndash;27.0 mg/g tobacco. (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e, Sl. No. 1; See Suppl. Information for further details). However, menthol could not be traced in smokeless tobacco products such as gul (number of samples, n\u0026thinsp;=\u0026thinsp;10), sada (n\u0026thinsp;=\u0026thinsp;3), gudakhu (n\u0026thinsp;=\u0026thinsp;7), snuff (n\u0026thinsp;=\u0026thinsp;3) and khaini (n\u0026thinsp;=\u0026thinsp;6) products (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e, Suppl. Information). Moreover, 3 brands of khaini and sada samples contain trace level (0.23\u0026ndash;0.34 mg/g) of menthol in it. 8 brands of khaini \u0026amp; sada products such as Raja Tobacco, Afgan Rally, Chunnu Right Choice, Golden Sada etc. and 2 brands of snus products such as Chaini khaini, Bolt Filter contain menthol as an additive (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). The amount of menthol in these 8 brands of khaini \u0026amp; sada products were present in the range of 1.5\u0026ndash;8.7 mg/g tobacco (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). Two kiwam products were tested and menthol was present in both the samples in the range of 13.0\u0026ndash;24.9 mg/g tobacco.\u003c/p\u003e \u003cp\u003eThe laboratory has quantified the menthol content in various cigarette and bidi samples (Tables\u0026nbsp;3 and 4, Suppl. Information). Some cigarette brands such as Gold Flake Indie Mint, Wave Cool Mint, Total, Wave Fruit Mint, Topaz, Players contain menthol ball/capsule\u003csup\u003e33,34\u003c/sup\u003e inside the cigarette filter which are broken by pressing the filter of these types of cigarettes during smoking by the consumer.\u003c/p\u003e \u003cp\u003eMenthol was estimated in cigarette as mg/cigarette as two whole cigarette were taken for analysis as per the CORESTA standard method. The laboratory analyzed 28 numbers of different brands of cigarettes and among them 11 brands of cigarette contain menthol in the range of 0.02\u0026ndash;9.3 mg per cigarette (Fig.\u0026nbsp;2). It indicates that around 39% of cigarette samples sold in Indian market contain menthol as per the present study is concerned. Our laboratory has tested 15 number of different brands of bidi samples and menthol is found in the range of ND \u0026ndash; 0.21 mg/g of tobacco filler of bidi sample (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e; Sl. No. 11).\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eMenthol content in different brands of smokeless tobacco samples and smoke tobacco samples:\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"6\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSl. No.\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eType of Tobacco Sample\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSmokeless / Smoke Tobacco\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eNo. of Brands analyzed\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eMenthol\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003eUnit\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eChewing Tobacco \u0026amp; Zarda\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSmokeless\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e35\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e1.6\u0026ndash;27.0\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003emg/g\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSnus\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSmokeless\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e7.6\u0026ndash;8.3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003emg/g\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eKhaini\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSmokeless\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e14\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eND \u0026ndash; 8.7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003emg/g\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSada\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSmokeless\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e6\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eND \u0026ndash; 3.5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003emg/g\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eGul\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSmokeless\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e10\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eND\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003emg/g\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e6\u003csup\u003e#\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eDried leaf\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSmokeless\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eND\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003emg/g\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eGurakhu\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSmokeless\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eND\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003emg/g\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSnuff\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSmokeless\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eND\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003emg/g\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e9\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eKiwam\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSmokeless\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e13.0\u0026ndash;24.9\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003emg/g\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e10\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eCigarettes\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSmoke\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e28\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eND \u0026ndash; 9.3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003emg/cig.\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e11\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eBidi\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSmoke\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e15\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eND \u0026ndash; 0.21\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003emg/g\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eND: Not Detected. (Limit of Detection of Menthol Glycerol\u0026thinsp;=\u0026thinsp;0.013 mg/g). # Received from different sources.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eHumectant (Glycerol and 1,2-Propylene Glycol)\u003c/h3\u003e\n\u003cp\u003eThe most commonly used humectants in tobacco products are glycerol and 1,2-propylene glycol.\u003csup\u003e\u003cspan additionalcitationids=\"CR24 CR25\" citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e\u003c/sup\u003e This laboratory has tested a variety of smokeless tobacco products and among them zarda, chewing tobacco and kiwam contain high levels of glycerol and 1,2 \u0026ndash; propylene glycol. Mean glycerol and 1,2- propylene glycol were found in the range of 0.9\u0026ndash;185.3 mg/g and ND \u0026ndash; 40.8 mg/g respectively (Table\u0026nbsp;\u003cspan refid=\"Tab2\" class=\"InternalRef\"\u003e2\u003c/span\u003e; See Table\u0026nbsp;5 Suppl. Information for more details)). A kiwam samples Raj Ratan tobacco contain exceptionally high quantity of glycerol (456.28 mg/g tobacco) whereas Royal Tulsi contain both glycerol (2.3 mg/g tobacco) and 1,2-propylene glycol (23.9 mg/g tobacco) (Table\u0026nbsp;6, Suppl. Information). 10 number of gul samples were analysed for the presence and quantification of humectants (Table\u0026nbsp;6, Suppl. Information). Among them glycerol was present in 1 gul samples (1.12 mg/g) whereas another 1 gul sample contain 1.11 mg/g of 1,2-propylene glycol. Other smokeless tobacco products such as snuff, gurakhu, dried leaf and sada do not contain any glycerol and 1,2-propylene glycol (Table\u0026nbsp;6, Suppl. Information). Among 14 khaini samples tested in this laboratory, 7 samples do not contain glycerol, 4 samples contain trace quantity of glycerol which is below the limit of quantitation and 3 samples contain glycerol in the range of 1.23\u0026ndash;4.56 mg/g tobacco (Table\u0026nbsp;6, Suppl. Information). However among 14 khaini samples 1,2 \u0026ndash; propylene glycol was absent in 13 khaini samples and 1 khaini sample contain 0.64 mg/g tobacco.\u003c/p\u003e \u003cp\u003e \u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab2\" border=\"1\"\u003e \u003ccaption language=\"En\"\u003e \u003cdiv class=\"CaptionNumber\"\u003eTable 2\u003c/div\u003e \u003cdiv class=\"CaptionContent\"\u003e \u003cp\u003eDetermination of Additives (Glycerol and 1,2-Propylene Glycol) in different variety of smokeless and smoke tobacco samples.\u003c/p\u003e \u003c/div\u003e \u003c/caption\u003e \u003ccolgroup cols=\"6\"\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e \u003cdiv align=\"char\" char=\".\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e \u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e \u003cthead\u003e \u003ctr\u003e \u003cth align=\"left\" colname=\"c1\"\u003e \u003cp\u003eSl. No.\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c2\"\u003e \u003cp\u003eTobacco Sample\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c3\"\u003e \u003cp\u003eTypes of Tobacco Sample\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c4\"\u003e \u003cp\u003eNo. of Brands analyzed\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c5\"\u003e \u003cp\u003eGlycerol (mg/g)\u003c/p\u003e \u003c/th\u003e \u003cth align=\"left\" colname=\"c6\"\u003e \u003cp\u003ePG (mg/g)\u003c/p\u003e \u003c/th\u003e \u003c/tr\u003e \u003c/thead\u003e \u003ctbody\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e1\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eChewing Tobacco \u0026amp; Zarda\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSLT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e35\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e0.9\u0026ndash;185.3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eND \u0026ndash; 40.8\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSnus\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSLT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e39.5\u0026ndash;53.4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003e2.1\u0026ndash;16.6\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eGul\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSLT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e10\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eND \u0026ndash; 1.12\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eND \u0026minus;\u0026thinsp;1.11\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e4\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eKhaini\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSLT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e14\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eND \u0026ndash; 4.56\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eND \u0026ndash; 0.64\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSada\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSLT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e6\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eND\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eND\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e6\u003csup\u003e#\u003c/sup\u003e\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eDried leaf\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSLT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e5\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eND\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eND\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e7\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eGurakhu\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSLT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eND\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eND\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e8\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eSnuff\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSLT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eND\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eND\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e9\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eKiwam\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSLT\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e2\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003e2.26\u0026ndash;456.3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eND \u0026ndash; 23.9\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e10\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eCigarette\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSmoke\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e28\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eND \u0026minus;\u0026thinsp;22.3\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eND \u0026ndash; 10.7\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003ctr\u003e \u003ctd align=\"left\" colname=\"c1\"\u003e \u003cp\u003e11\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c2\"\u003e \u003cp\u003eBidi\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c3\"\u003e \u003cp\u003eSmoke\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"char\" char=\".\" colname=\"c4\"\u003e \u003cp\u003e15\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c5\"\u003e \u003cp\u003eND \u0026minus;\u0026thinsp;3.6\u003c/p\u003e \u003c/td\u003e \u003ctd align=\"left\" colname=\"c6\"\u003e \u003cp\u003eND\u003c/p\u003e \u003c/td\u003e \u003c/tr\u003e \u003c/tbody\u003e \u003c/colgroup\u003e \u003c/table\u003e\u003c/div\u003e \u003c/p\u003e \u003cp\u003eND: Not Detected. BLOQ: below limit of quantitation. (Limit of Quantitation of Glycerol\u0026thinsp;=\u0026thinsp;0.9 mg/g and 1,2-Propylene Glycol\u0026thinsp;=\u0026thinsp;0.2 mg/g). # Received from different sources.\u003c/p\u003e \u003cp\u003eAmong smoke tobacco samples, bidi and cigarettes samples were analyzed for the quantification of glycerol and 1,2-propylene glycol (PG). Out of 28 cigarette samples tested, 27 number of cigarette samples contain glycerol or 1,2-propylene glycol as one of the additives. The amount of glycerol and 1,2-propylene glycol was present in cigarette sample ranges from not detected (ND) to 22.33 mg/g tobacco (Mean\u0026thinsp;=\u0026thinsp;10.2 mg/g) and ND to 10.70 mg/g tobacco (Mean\u0026thinsp;=\u0026thinsp;2.2 mg/g) respectively (Table\u0026nbsp;7, Suppl. Information). Among the cigarette samples tested, only 4 cigarette samples P-10 premium, Charms, H-10 and Sunils Filter did not contain glycerol as an additive whereas 3 samples did not contain 1,2-propylene glycol and 8 samples contain trace quantity of 1,2-propylene glycol which is below limit of quantitation.\u003c/p\u003e \u003cp\u003eAmong the 15 bidi brands tested in our laboratory, 1,2-propylene glycol was not detected in all samples whereas glycerol was not present in 7 samples. 8 bidi samples contain glycerol in the range of 1.1\u0026ndash;3.6 mg/g (Mean\u0026thinsp;=\u0026thinsp;1.9 mg/g tobacco).\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eSmokeless tobacco products sold in India contain high levels of flavors and sweetening agent to enhance the palatability of tobacco product. Among the flavors menthol is most extensively used chemicals in tobacco products due to their cooling effect which help to mask the harshness of the products.\u003csup\u003e\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e,\u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e,\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e\u003c/sup\u003e Lisko \u003cem\u003eet al.\u003c/em\u003e in 2014 studied the levels of flavors in smokeless tobacco products of South East Asia of varieties like zarda, quiwam, gutkha, and khaini samples where menthol was present in zarda samples in the range 5\u0026ndash;22 mg/g tobacco.\u003csup\u003e\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e\u003c/sup\u003e Cheah and coworkers recently carried out the study of analysis of composition of smokeless tobacco products (zarda/chewing tobacco, khaini, kharra and gudakhu) sold in north, east west and central regions of India. Menthol was measured in two zarda samples and found in the range of 4.1\u0026ndash;9.7 mg/g tobacco whereas in three chewing tobacco, menthol was found in the range of 0.3\u0026ndash;2.0 mg/g tobacco.\u003csup\u003e\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e\u003c/sup\u003e\u003c/p\u003e \u003cp\u003eIn the present study, menthol was detected in all the zarda and chewing tobacco samples and was found in the range of 1.6\u0026ndash;27.0 mg/g tobacco, which is consistent with the findings of Lisko \u0026amp; coworkers.\u003csup\u003e\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e\u003c/sup\u003e In 2018, Stanfill \u003cem\u003eet al.\u003c/em\u003e investigated the composition of packaged SLT products available in a well-established South Asian community outside of South Asia. In their work menthol content was reported in the range of 0.16\u0026ndash;21.7 mg/g tobacco and 7.0\u0026ndash;11.0 mg/g tobacco respectively for zarda and khaini samples which are comparable to the present study where samples are mostly collected from North-Eastern part of India.\u003csup\u003e\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e\u003c/sup\u003e From the observations, it can be inferred that smokeless tobacco products sold in Indian market contains varying menthol concentrations; upto 17 fold in zarda \u0026amp; chewing tobacco from lowest quantity quantified in zarda \u0026amp; chewing tobacco sample. In our investigation, the snus products Chaini khaini, Bolt Filter contain menthol in the range of 7.6\u0026ndash;8.3 mg/g tobacco. Stepanov \u003cem\u003eet al.\u003c/em\u003e mentioned in their brief report that Chaini khaini is marketed as a safe alternative to other tobacco products despite of containing high levels of carcinogenic nitrosamines and biologically available nicotine.\u003csup\u003e\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e\u003c/sup\u003e Stanfill and coworkers investigated the menthol content in four kiwam products and contain menthol (0.85\u0026ndash;12.3 mg/g tobacco) in all products.\u003csup\u003e\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e\u003c/sup\u003e In our present study, two samples were investigated which contain menthol (13.0-24.9 mg/g tobacco). The consumption of such mentholated flavored tobacco may enhance the addiction towards such harmful products and regulating or banning menthol additives should have immediate focus for better public health.\u003c/p\u003e \u003cp\u003eIn the present study, the menthol content in mentholated cigarettes was found in the range of 2.3\u0026ndash;9.3 mg/cigarette (Table\u0026nbsp;3, Suppl. Information). Taylor \u003cem\u003eet al\u003c/em\u003e. has studied the menthol content in US marketed cigarettes and found that menthol content in 23 menthol branded cigarettes in the range of 2.9\u0026ndash;19.6 mg/cigarette whereas in non-mentholated cigarette menthol was present in the range of 0.002\u0026ndash;0.07 mg/cigarette.\u003csup\u003e38\u003c/sup\u003e The observations of our laboratory were consistent with the findings of Taylor \u0026amp; coworkers but highest menthol content in mentholated Indian cigarette is 9.3 mg/cigarette which is somewhat lesser than US marketed cigarettes which is 19.6 mg/cigarette. Silva \u003cem\u003eet al.\u003c/em\u003e reported in their article that tobacco companies use menthol in 'non-menthol' cigarettes at subliminal levels, i.e., at levels not perceptible as menthol-flavored.\u003csup\u003e\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e\u003c/sup\u003e In this investigation, in two non-mentholated cigarettes viz. silk cut and gold flake premium, menthol was found in the range of 0.02\u0026ndash;0.07 mg/cigarette (Fig.\u0026nbsp;2) which are at subliminal level.\u003c/p\u003e \u003cp\u003eWu group mentioned that bidi cigarettes sold in the USA contain various flavorings, including vanilla, chocolate, strawberry, grape, root beer and others whereas Indian bidis do not contain any flavor additives.\u003csup\u003e\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e,\u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e\u003c/sup\u003e Stanfill \u0026amp; coworkers reported that menthol was present in the range of ND \u0026ndash; 2.77 mg/bidi in 10 of the flavored bidi brands including non-mentholated brands.\u003csup\u003e\u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e\u003c/sup\u003e Our laboratory has tested 15 number of different brands of bidi samples where menthol could not be traced in 11 samples (Table\u0026nbsp;4, Suppl. Information). In two bidi samples menthol was present in the range of 0.15\u0026ndash;0.21 mg/g tobacco whereas in another two samples menthol was detected which was below limit of quantification. (LOQ\u0026thinsp;=\u0026thinsp;0.04 mg/g). From this studies, it can be inferred that most Indian bidis do not contain menthol additives.\u003c/p\u003e \u003cp\u003eThe additives glycerol and 1,2-propylene glycol, used as humectants in tobacco products, contain varying levels of these substances. Gupta \u003cem\u003eet al\u003c/em\u003e. in their report mentioned that the chewing tobacco products contain high levels of humectants, with mean glycerol levels between 24 to 40 mg/g tobacco and propylene glycol between 1.9 to 8.0 mg/g tobacco.\u003csup\u003e\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e\u003c/sup\u003e In our findings seven chewing tobacco products and zarda contain exceptionally high quantities of glycerol around 2\u0026ndash;5 fold higher than the high value of humectants found in most of the chewing tobacco products (Table\u0026nbsp;5, Sl. No. 29\u0026ndash;35, Supplementary Information). Foulds \u003cem\u003eet al.\u003c/em\u003e and Rickert \u003cem\u003eet al.\u003c/em\u003e reported that Swedish snus contains 15\u0026ndash;35 mg/g humectants (e.g. propylene glycol and glycerol) and 16.2\u0026ndash;16.4 mg/g 1,2-propylene glycol resepectively.\u003csup\u003e\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e,\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e\u003c/sup\u003e In our investigation snus products such as Chaini filter pouch, Bolt Filter contain glycerol and 1,2 \u0026ndash; propylene glycol in the range of 39.5\u0026ndash;53.4 mg/g tobacco and 2.1\u0026ndash;16.6 tobacco mg/g respectively (Table\u0026nbsp;6, Supplementary Information). Rickert \u003cem\u003eet al\u003c/em\u003e. studied the presence of glycerol and 1,2-propylene glycol in 25 brands of snuff products and found that glycerol was absent in all products whereas 1,2-propylene glycol was present in only 7 samples in the range of 3.92\u0026ndash;23.4 mg/g tobacco.\u003csup\u003e\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e\u003c/sup\u003e However, in our investigation all the three snuff products do not contain glycerol and 1,2-propylene glycol.\u003c/p\u003e \u003cp\u003eIn our findings the amount of glycerol and 1,2-propylene glycol in cigarette samples were present in the range of ND \u0026ndash; 22.33 mg/g tobacco and ND \u0026ndash; 10.70 mg/g tobacco. Pennings and coworkers measured the quantity of glycerol and 1,2-propylene glycol in 52 tobacco samples and found that these substances were present in the range of 1.1\u0026ndash;23.7 mg/g of tobacco (mean\u0026thinsp;=\u0026thinsp;7.8 mg/g) and 0\u0026ndash;14.8 mg/g of tobacco (mean\u0026thinsp;=\u0026thinsp;4.6 mg/g), respectively.\u003csup\u003e\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e\u003c/sup\u003e In our investigation, the maximum amount of 1,2-propylene glycol in cigarettes was found to be 10.7 mg/g tobacco which is lower than the findings of the Pennings group, i.e. 14.8 mg/g tobacco. It was also observed that glycerol was used as a humectant in most cigarettes more frequently than 1,2-propylene glycol. The amount of glycerol (1.06\u0026ndash;22.33 mg/g tobacco, mean\u0026thinsp;=\u0026thinsp;10.2 mg/g) added in cigarettes are also comparatively higher than 1,2-propylene glycol (0.3\u0026ndash;10.7 mg/g tobacco; mean\u0026thinsp;=\u0026thinsp;2.2 mg/g). Most cigarettes contain glycerol and/or 1,2-propylene glycol as additives, which makes the product more toxic, as these additives produce harmful aldehydes when burned. In comparison of cigarette and bidi products, cigarette products contain high quantity of glycerol almost 7 fold higher compared to bidi products (Table\u0026nbsp;8, Suppl. Information). Chewing tobacco/ zarda and kiwam products contain exceptionally high quantity of glycerol upto 18% and 45% respectively compared to the cigarettes which contain upto 2.2% glycerol.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eAccording to recommendations from the WHO TobReg, it is crucial to implement product standards and strategies to mandate the reduction of toxicants in all types of tobacco products by removing or lowering the contents such as menthol and humectant.\u003csup\u003e\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e,\u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e\u003c/sup\u003e The contents menthol, glycerol and 1,2-propylene glycol in smoke and smokeless tobacco products available in India are poorly studied and so far received limited attention from researchers and policy makers. The above findings may be useful to regulate the additives in the tobacco products to minimize the exposures and risks associated with the use of smoke and smokeless tobacco products.\u003c/p\u003e "},{"header":"Declarations","content":"\u003cp\u003e \u003ch2\u003eCompeting Interests\u003c/h2\u003e \u003cp\u003eThe authors declare no conflict of interests and no financial interests.\u003c/p\u003e \u003c/p\u003e\u003ch2\u003eAuthor Contribution\u003c/h2\u003e\u003cp\u003eThe experimental work was carried out by D.R.D., A.K.Y., S.K.S. and K.J.B. at NTTL Guwahati. The results were analyzed by P.S. The main manuscript text was prepared by P.S. and D.R.D. The manuscript text was reviewed and edited by D.K. The manuscript was reviewed by all authors\u003c/p\u003e\u003ch2\u003eAcknowledgement\u003c/h2\u003e\u003cp\u003eThe finding and conclusions reported in this article are those of the author(s) and do not necessarily represent the views of the National Tobacco Control Program (NTCP), Ministry of health \u0026amp; Family Welfare, Government of India. Brand names are used for identification only and the laboratory does not endorse any tobacco product. We greatly acknowledge Dr. L. Swasticharan, Addl. DDG, NTCP, State Nodal Officers, NTCP and National Tobacco Control Program, Ministry of Health \u0026amp; Family Welfare, Government of India for the support in this investigation.\u003c/p\u003e\u003ch2\u003eData Availability\u003c/h2\u003e\u003cp\u003eThe data generated are incorporated into the article and its online supplementary material.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eTata Institute of Social Sciences (TISS), Mumbai and Ministry of Health and Family Welfare, Government of India. Global Adult Tobacco Survey GATS 2 India 2016-17. http://download.tiss.edu/Global_Adult_Tobacco_Survey2_India_2016-17_June2018.pdf\u003c/li\u003e\n\u003cli\u003eHealth Topics Tobacco \u0026ndash; World Health Organization. https://www.who.int/india/health-topics/tobacco. (Accessed January 2024).\u003c/li\u003e\n\u003cli\u003ePennings, J. L. A., Cremers, J. W. J. M., Becker, M. J. A., Klerx, W. N. M. \u0026amp; Talhout, R. 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Impact assessment of WHO TobReg proposals for mandated lowering of selected mainstream cigarette smoke toxicants. \u003cem\u003eRegul. Toxicol. Pharmacol.\u003c/em\u003e\u003cstrong\u003e86\u003c/strong\u003e, 332-348 (2017).\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"","lastPublishedDoi":"10.21203/rs.3.rs-6161281/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-6161281/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eDifferent chemicals are added to smoke and smokeless tobacco products to make the products more attractive and palatable. In this article, 43 smoke and 84 smokeless tobacco products were investigated to detect and quantify menthol, glycerol, and 1,2-propylene glycol. The data obtained were compared across the tobacco products to understand variation of these chemicals in different categories of tobacco products. The smokeless tobacco products zarda, chewing tobacco, khaini and snus contain menthol as an additive. Some smokeless tobacco such as chewing tobacco, zarda, snus and kiwam contain glycerol and / or 1,2-propylene glycol as additives. One smokeless tobacco products contain 45% glycerol which is exceptionally high compared to reported glycerol level in other studies. Among the tested 28 number of cigarette samples 39% of cigarette samples contain menthol as an additive. Most cigarettes contain both glycerol or 1,2-propylene glycol or one of them as humectants whereas bidi do not contain menthol and 1,2\u0026ndash;propylene glycol. The determination of these additives present in the tobacco products may be helpful in further understanding the health hazards associated with these products for tobacco users. The findings in this work may provide valuable information / data to regulate the additive content in tobacco products for effective tobacco control in India.\u003c/p\u003e","manuscriptTitle":"Quantitative Estimation of Menthol, Glycerol and 1,2-Propylene Glycol in Tobacco Products Available in India","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-04-21 12:09:10","doi":"10.21203/rs.3.rs-6161281/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"9b4f5e3f-7d8a-471c-a8cc-594d60afe3d3","owner":[],"postedDate":"April 21st, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":47207855,"name":"Health sciences/Medical research"},{"id":47207856,"name":"Physical sciences/Chemistry"}],"tags":[],"updatedAt":"2026-02-17T09:11:31+00:00","versionOfRecord":[],"versionCreatedAt":"2025-04-21 12:09:10","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-6161281","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-6161281","identity":"rs-6161281","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}