12.5 Measuring emissions and exposure to tobacco products

Last updated: January 2022
Suggested citation: Winnall, WR. 12.5 Measuring emissions and exposure to tobacco products. In Greenhalgh EM, Scollo, MM and Winstanley, MH [editors]. Tobacco in Australia: Facts and issues. Melbourne: Cancer Council Victoria; 2022. Available from https://www.tobaccoinaustralia.org.au/chapter-12-tobacco-products/12-5-measuring-emissions-and-exposure-to-tobacco-products  


Tobacco emissions are all the chemicals released from a tobacco product when used as intended. These include the chemicals in smoke from cigarettes, cigars, pipes, waterpipes, bidis and kreteks, as well as the emissions from heated tobacco products and smokeless tobacco. Tobacco emissions cause innumerable health effects as well as addiction to smoking and the deaths of millions of people each year. The chemicals that make up these emissions are therefore subject to scientific research, industry reporting, and regulation by governments that are attempting to reduce the health impacts of smoking.

This section discusses the methods used for measuring the chemicals that constitute tobacco product emissions and the use of biomarkers to understand human exposure. The chemicals present in tobacco before burning are discussed in Section 12.3. Many new chemicals are produced during the burning of tobacco, some of which have carcinogenic (cancer-causing) and other toxic activity, as are discussed in Section 12.4.

Tobacco companies may measure emissions for quality control, research and development, and mandatory reporting. Researchers measure emissions to determine which are present and what their effects may be on humans and on the environment. Regulators, acting under advice from the WHO, use these methods for quantifying emissions to facilitate and regulate the mandatory reporting of these chemicals, with the ultimate aim of reducing the harmful effects of tobacco emissions.

Understanding the effects of the chemicals in tobacco emissions also requires knowledge of which chemicals enter the body and to which parts of the body they move. The use of biomarkers to study human exposure is increasingly improving knowledge in this area, as described below in Section 12.5.6.

12.5.1 Measuring tar and emissions from cigarettes using smoking machines

Measuring the amount of chemicals in cigarette smoke involves laboratory techniques that have evolved and improved over time. Smoking machines have been used for decades to compare the tar, nicotine and carbon monoxide in mainstream smoke from one brand of cigarette to another under standardised conditions. For many years, the results from these measurements were erroneously assumed to correlate with the degree to which these cigarettes are harming human health. In some regions, these results have served as regulatory standards, where brands that generate emissions above a set limit for tar, nicotine and carbon monoxide were prohibited.1 However, there are multiple problems with this approach, making it inadequate for regulation and consumer health information.

Smoking machines physically separate the smoke into a gaseous phase (“vapour phase”) and a particle phase, from which individual chemicals can be detected. The particle phase contains nicotine, some water, and most of the chemicals from the aerosol particles of smoke (commonly referred to as ‘tar’).

A smoking machine method to compare the amount of tar elicited from different types of cigarettes was developed by the Federal Trade Commission (FTC) in the US in 1967. The aim, from the outset, was to obtain standardised data about the amount of tar and nicotine in mainstream smoke, not to replicate the actions of human smokers.2 The FTC method was a modified form of the earlier Cambridge filter method.3 A smoking machine with one or multiple openings is used to “smoke” the cigarettes using suction. A filter pad is used to collect the ‘particle phase’ and gases are separately collected for analysis. Under arbitrary standard conditions used by the FTC from 1967, cigarettes are smoked using 1 puff (duration of 2 seconds and 35 ml volume) every minute, with butt length of 23 mm (or the length of the overwrap plus 3 mm, whichever is longer). From the residue collected on the filter pad the amount of nicotine is measured, and the amount of tar is calculated from the weight of the residue on the pad minus the nicotine and estimated water content. Carbon monoxide levels are measured in the isolated gas fraction.2

In 2008, the FTC rescinded their guidance from the 1960s on the use of the FTC method for measuring tar and nicotine by smoking machines.4 See Section 12.5.2 for more details.

The International Organization for Standardization (ISO) also developed a smoking machine method to measure tar, nicotine and carbon monoxide levels in cigarette smoke.5 The method developed by the ISO is very similar to the FTC measure but is done at a slightly different temperature and cigarette butt length.5 ,6

The tar collected from smoking machines includes many of the toxic and addictive chemicals from cigarette smoke. When the tar yield of a product is stated on a cigarette packet, as was the case in Australia until 2006—and as is still the case in many countries—it doesn’t include the amount of water and nicotine in the residue. ‘Tar’ is also used colloquially to refer to the brown stains seen on the end of cigarette filters and on smokers’ fingers, and to the build-up of chemicals as a sticky layer in the lungs of smokers.7

In addition to tar, smoking machines and other methods were also used to detect nicotine, the primary addictive ingredient of tobacco smoke, and carbon monoxide, due to it being a likely cause of cardiovascular damage.

In the 1950s it was shown that tumours developed when tar dissolved in acetone was painted onto mouse skin.8 ,9 There was a dose–response relationship between the amount of tar to which mice were exposed and the frequency with which tumours developed in mice.10 ,11 It was assumed that this would also be true for humans, and that reducing smokers’ exposure to tar could reduce the risk of tumours in smokers. This led to efforts to reduce the amount of tar to which smokers are exposed and the measuring and reporting of the amount of tar elicited by different types of cigarettes. However, we now know that the tar yields measured by smoking machines are not associated with reduced risk of cancers in smokers. See Section 12.5.2 below for more details.

Since the 1990s, the smoking machine methods have been varied by using different conditions in an attempt to emulate different types of smoking by consumers. The ISO Intensive Condition test (also called the Canadian regime) is the most notable. During this protocol, the machine takes a 55ml puff of 2 seconds duration, once every 30 seconds, with all filter ventilation holes blocked (see Sections and 12.5.2 below for more details). This method was recommended by the WHO TobReg committee in 2004 as it comes close to representing the maximum exposure level to which an ordinary smoker could reasonably be expected to be subject when smoking.12 When a range of cigarette brands were tested in Canada using the ISO Intensive Condition test, over double the amount of nicotine was measured on average, compared to the standard ISO conditions.

12.5.2 Compensatory smoking and other issues with smoking machine methods

For many years the ISO or FTC estimated yields of tar, nicotine and carbon monoxide were stated for specific cigarette brands. They were included on the packaging of Australian cigarettes until 2006. These values have given a false impression that some brands of cigarettes would reduce the risk to smokers. The accuracy of these yields in predicting human exposure is now known to be quite poor.

The most serious problem with the ISO and FTC machine smoking methods is that the smoking parameters systematically underestimate the intensity of people’s smoking behaviour.1 These methods assume that individual smokers will always take the same volume of smoke from any cigarette. For most addicted smokers, their target from smoking is nicotine intake and not the volume of smoke they take from each cigarette.13 The majority of addicted smokers appear to require somewhere between 0.9 mg and 1.4 mg of nicotine from each cigarette for it to be satisfying.14 Smokers subconsciously change smoking parameters such as puff size and time taken between puffs when they change brands, in order to achieve their target nicotine intakes.15 ,16These changes in smoking behaviour are known as compensatory smoking (also discussed in Sections and As reported tar and nicotine yields decrease (when smokers move to brands with lower reported yields) parameters such as puff size and total number of puffs taken per cigarette increase. Attempting to account for more intensive smoking, the ISO intensive condition test uses larger volume puffs at a greater frequency. However, this test was not better at predicting nicotine uptake, when compared to a salivary biomarker measured in smokers.1  

An important determinant of nicotine levels measured by the FTC and ISO standard methods are ventilation holes in the cigarette filter. These filter ventilation holes dilute the smoke with air, as described in Section Smokers, however, partially cover the ventilation holes with their fingers and lips, as well as smoking more intensely. By producing cigarettes modified for filter ventilation, the industry has made a product that performs differently under standard testing conditions than it does when used by consumers. The ISO Intensive Condition test requires complete covering of these holes to attempt to correct for the differences caused by filter ventilation holes.17

The standard FTC and ISO methods exaggerate the differences between brands, falsely indicating that some brands are a lower risk to smokers.1 Cigarette brands have been described as “regular”, “light” or “ultralight” based on their tar levels. However, people who smoked “light” or “ultralight” brands did not have a lower risk of lung cancer compared to smokers of regular cigarettes. Experiments have failed to find an association between stated yields of tar and biomarkers of exposure in humans.18 The tobacco industry has modified its products to produce low levels of tar, nicotine and carbon monoxide during the machine tests, but smokers themselves received little to no reduction in levels.1

Another problem with the tar yield is that tar varies significantly in composition between different cigarette brands or tobacco varieties.19 ,20There is evidence that concentrations in smoke of certain known carcinogens vary substantially between cigarette types and brands. For example, smoke from cigarettes made with the Burley tobacco variety have been found to have considerably higher levels of carcinogenic N'-nitrosonornicotine (NNN) compared to those made with Virginia or Oriental tobacco (1,970 ng/g Burley; 35 ng/g Virginia and 84 ng/g Oriental).21 Carcinogenic 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) was also considerably higher in cigarettes made from Burley tobacco.21 The proportion of Burley tobacco in cigarettes made from blended tobacco is likely to affect the amount of NNN and NNK in their smoke. Data from Philip Morris from 2005 have shown a wide range in these chemicals found in emissions from its cigarettes. In the brands of cigarettes tested at the time, the amount of NNN ranged from 16 to 189 ng/mg and NNK from 23 to 111 ng/mg.22 However, there is no evidence that any form of tar with lower amounts of one carcinogen leads to better outcomes in smokers.

Rather than measure tar as a whole to infer the potential for harm, the current best practice focusses on measuring individual toxicants in tobacco emissions, as described below.

12.5.3 Development of best practice in measuring chemicals in mainstream cigarette emissions

TobLabNet (Tobacco Laboratory Network, under the World Health Organization) is a global network of government, academic, and independent laboratories that collaborate to strengthen capacity for the testing of tobacco product contents and emissions. TobLabNet has produced a list of 39 chemicals in cigarette smoke that should be measured to monitor tobacco products in each country.23 In the US, the FDA has also produced a list of 93 harmful and potentially harmful constituents in tobacco products and smoke.24 For the purposes of this section, the recommendations by TobLabNet are considered current best practice in the detection of specific toxic chemicals in cigarette smoke. See Section 12.4.3 for more details on the chemicals discussed here.

Detection of chemicals in mainstream cigarette smoke requires the generation of smoke, making it inevitable that machine smoking methods must still be used during detection of these chemicals. The current recommendations from TobLabNet use smoking machines with the ISO Standard Conditions (35 ml puffs every 60 seconds) and the ISO Intensive Condition test (55ml puff every 30 seconds with filter ventilation blocked).25 Use of smoking machines, however, is of limited value for inferring human health risks. The WHO clarify this in its statement:

‘No machine smoking regimen can represent all human smoking behaviour: machine smoking testing is useful for characterizing cigarette emissions for design and regulatory purposes, but communication of machine measurements to smokers can result in misunderstanding about differences between brands in exposure and risk. Data on smoke emissions from machine measurements may be used as inputs for product hazard assessment, but they are not intended to be nor are they valid as measures of human exposure or risks. Representing differences in machine measurements as differences in exposure or risk is a misuse of testing with WHO TobLabNet standards.’ 25

This is reiterated in the findings of the 2010 US Surgeon General’s report that:

‘There is no available cigarette machine-smoking method that can be used to accurately predict doses of the chemical constituents of tobacco smoke received when using tobacco products.’

The WHO recommends moving towards using limits for certain smoke chemicals expressed per mg of nicotine rather than per cigarette. Its reasoning is that people smoke cigarettes to obtain adequate nicotine to satisfy their physiological and behavioural needs, and most toxic chemicals in smoke are delivered in proportion to nicotine.23 ,26 The chemicals produced by smoking under standardised conditions (such as in smoking machine experiments) are useful in establishing the data needed to establish these limits.

As of 2018, TobLabNet scientists had so far developed and validated standard operative procedures to determine the amounts of 12 of the 39 chemicals in cigarette smoke recommended for monitoring in every country. All these methods use smoking machines to separate the phases and other laboratory methods for detection and quantification of each specific chemical.23 Each of these chemicals and methods are listed in Table 12.5.1 and the detection methods described briefly below:

HPLC (High-Performance Liquid Chromatography) HPLC uses a pump to force a mix of chemicals onto an adsorbent material. Each chemical binds and is released from the adsorbent material at different rates, based on its particular properties. This separates the mix into groups of chemicals, allowing for the measurement of individual chemicals.27

Mass spectrometry During mass spectrometry, chemicals in a mix are fragmented and the fragments are separated in an electric field based on their mass and electric charge. Analysis of the fragmentation pattern can determine the structure of the original chemicals. Mass spectrometry can also determine how much of a specific chemical is in a mixture.27

Gas chromatography Gas chromatography is similar to HPLC, except that the mixture of chemicals is converted into a gas prior to binding and release from the adsorbent material. Gas chromatography can be used to determine the amount of specific chemicals in a mix.27

Non-dispersive infrared (NDIR) During NDIR analysis, infrared light is directed through a mix of chemicals. A chemical, such as carbon monoxide, will absorb the light at a specific wavelength, which can be measured to identify and quantitate (very precisely quantify) the amount in the mix.28

In addition to the standard operating procedures and chemicals described in Table 12.4.2, TobLabNet refers to scientific publications describing possible methods for the detection and quantitation of most of the remaining chemicals in its list of 39 to be measured.23 These are:

  • Aldehydes (butyraldehyde, crotonaldehyde, propionaldehyde)29 ,30
  • Aromatic amines (1-aminonaphthalene, 2-aminonaphthalene, 3-aminobiphenyl, 4-aminobiphenyl)31 ,32
  • Hydrocarbons (isoprene, toluene)33
  • Phenols (catechol, m-, p-, o-cresols, phenol, hydroquinone, resorcinol)34 ,35
  • Other organic compounds (acetone, acrylonitrile, pyridine, quinoline)29 ,33 ,36 ,37
  • Metals and metalloids (arsenic, cadmium, lead, mercury)38 ,39
  • Other constituents (hydrogen cyanide)40


Table 12.5.1 Chemicals measured in smoke emissions described by WHO standard operating procedures




Measurement method


3-(1-nitrosopyrrolidin-2-yl)-pyridine (NNN)

Particle phase


HPLC and mass spectrometry


SOP 03

4-(methylnitrosamino)-1-(3-pyridyl)-1 butanone (NNK)

Particle phase


HPLC and mass spectrometry


SOP 03

N-nitrosoanatabine (NAT)

Particle phase


HPLC and mass spectrometry


SOP 03

N-nitrosoanabasine (NAB)

Particle phase


HPLC and mass spectrometry


SOP 03


Particle phase


Gas chromatography and mass spectrometry

SOP 05


Particle phase

Carcinogen + toxic


SOP 08

acrolein (acrylaldehyde)

Particle phase



SOP 08


Particle phase



SOP 08



Particle phase

Carcinogen + toxic

Gas chromatography and mass spectrometry

SOP 09


Particle phase

Carcinogen + toxic

Gas chromatography and mass spectrometry

SOP 09


Particle phase


Gas chromatography

SOP 10

Carbon monoxide

Gas phase


Non-dispersive infrared analyser

SOP 10

Sources:  Report on the Scientific Basis of Tobacco Product Regulation: Seventh Report of a WHO Study Group,23 SOP 03,41 SOP 05,42 SOP 08,43 SOP 09,44 SOP 10.45


No methods had been recommended for measuring nitric oxides as of 2018.23 See Section 12.4.3 for more information about these chemicals.

12.5.4 Measuring emissions in sidestream smoke, secondhand and thirdhand cigarette smoke

Sidestream smoke emanates from the lit end of a cigarette between puffs. It is unfiltered, and produced by combustion and pyrolysis occurring at lower temperatures and lower oxygen levels than the conditions for mainstream smoke (see Section

Secondhand smoke consists of sidestream smoke, exhaled mainstream smoke, some mainstream smoke that escapes from the mouthpiece and any that passes across the porous paper wrapper (see Section Secondhand smoke is rapidly diluted as it dissipates into the air surrounding the cigarette. It undergoes changes in chemical content as it ages. The chemicals in secondhand smoke interact with other gases and particulate matter in the air around the passive smoker. Unlike mainstream smoke that reaches the lungs very rapidly after generation, secondhand smoke that people (including smokers themselves) breathe in during passive smoking is much more variable due to the effects of aging, dilution and interaction with other particles in the air.46 These differences make the measurement and reporting of secondhand and sidestream smoke emissions challenging. This section discusses measuring the chemicals in secondhand smoke from cigarettes. For a discussion of measuring exposure to secondhand smoke in people, see Section 4.4, and secondhand smoke from non-cigarette tobacco products is discussed in Sections and 12.4.4.

Sidestream smoke emissions from cigarettes can be analysed in a fixed (but artificial) environment by using smoking machines similar to those described for mainstream smoke above (Section 12.5.1). The smoke from the lit end of a cigarette is captured in a sealed container and the presence of tar and other chemicals is measured using similar methods to that for mainstream smoke. Early experiments showed that secondhand smoke contains many of the same chemicals as mainstream smoke, such as nicotine, carbon monoxide, toxic metals,47 and carcinogenic compounds.48 ,49 Some toxic chemicals are present at higher levels in sidestream smoke produced in this manner compared to mainstream smoke. These include polycyclic aromatic hydrocarbons (PAHs), N-nitrosamines, toxic metals, aromatic amines, aldehydes and other toxic organic compounds.48 ,50-54 These levels may be higher in sidestream smoke due to the lack of a filter and the differing combustion and pyrolysis conditions. This is consistent with animal experiments showing that extracts of sidestream smoke have a greater potential for carcinogenicity than from mainstream smoke.55 ,56 However, the disadvantage of controlled environment studies such as these is that they don’t accurately emulate real life situations. The amount of chemicals inhaled by a person from the sidestream smoke (and secondhand smoke) depends on many factors, such as the distance from the cigarette, airflow and ventilation in the area and the effects of aging on the smoke.46

Studies of secondhand smoke emissions, of which sidestream smoke makes up a considerable proportion, involve constructing scenarios that emulate conditions in which non-smokers inhale this smoke. Indoor smoking is commonly used as a model, involving exposure to higher levels of chemicals in secondhand smoke due to reduced ventilation compared to outdoor settings. An example is a study of secondhand smoke in houses; 90 to 200 minute old samples of secondhand smoke were collected from various sites inside three bedroom houses with poor ventilation, typical of those in cold climates.57 Nicotine and other chemicals from smoke were detected at each sampled site. Nicotine levels were lower in rooms further away from the smoking room.57 Similar studies have measured levels of toxic components of smoke in smoking rooms, nightclubs, taverns and living quarters, with considerably variability in the results.46 Chemicals detected in secondhand smoke from a range of studies include nicotine, carbon monoxide, hydrocarbons, aldehydes, PAHs and nitrosamines.46 ,58-61

Thirdhand smoke refers to the residual tobacco smoke constituents that remain on surfaces and in dust after tobacco has been smoked (see Section 4.3). A study of thirdhand cigarette smoke found evidence for the surface deposition of nicotine, nitrosamines and polycyclic aromatic hydrocarbons and de novo formation of NNK, on materials left in the vicinity of secondhand smoke for one hour.62 Research from Philip Morris showed that majority of the nicotine and tobacco-specific nitrosamines from indoor secondhand smoke remains on surfaces for months after smoking ceases.63 These experiments also indicated that the amount of NNK that persists months after smoking can exceed the amount that actually came out of the cigarettes,63 supporting the notion that this carcinogen is being produced from chemical reactions occurring in the thirdhand smoke residues.

12.5.5 Measuring emissions from other types of tobacco products

Measuring emissions from other types of tobacco products can be challenging and standard operating procedures are scarce. Taking these measurements requires knowledge of smokers’ puff topography parameters: such as the average puff number, volume and duration, and the inter-puff interval. These parameters are influential as they often strongly influence the concentrations of toxicant emissions. They may be difficult to determine or highly variable, making it challenging to define standardised smoking machine regimes. For some tobacco products, such as waterpipes, there is no exact time in which a smoking session is considered to have ended. Experimental measurement of chemicals in emissions from non-cigarette products that use non-standard conditions have produced highly variable results. Cigars

A cigar is a roll of tobacco that is wrapped in a tobacco leaf. There are three main types of cigars: large cigars, cigarillos and little cigars. With the exception of little cigars, cigars usually do not have a filter. Most cigar smokers only allow the smoke into their mouth and throat. However, people who smoke both cigars and cigarettes often inhale the cigar smoke into their lungs.64 See Section 12.2.1 for a description of the tobacco in cigars and Section 3.27.3 for the health effects of smoking cigars.

Cigars are made in a wide variety of sizes and shapes, ranging from little cigars that are similar to cigarettes, to large cigars that have up to 20 grams of tobacco and take up to 2 hours to smoke. The diameter of cigars ranges from about 6 mm to 25 mm.65 Puffing parameters also vary considerably between users, such as total puff volumes ranging from 84 to 732 ml for little cigars and 270 to 2089 ml for cigarillos, determined in one study.66 The variety in sizes and shapes of cigars and the smoking habits of their users complicates the measurement and comparisons of emissions from different types of cigars.

Cigar emissions can be analysed after being collected by cigarette smoking machines, which have been adapted to fit the size of the cigar. The International Organization for Standardization and the WHO’s Tobacco Product Regulation Study Group do not have any standard operating procedures for the analysis of cigar emissions using smoking machines. However, the tobacco industry association Cooperation Centre for Scientific Research Relative to Tobacco (CORESTA) have published a procedure for the analysis of non-hand-made cigar emissions by smoking machines.65

Given their similar size to cigarettes, the emissions of little cigars have been studied using smoking machines designed for cigarettes.66 ,67 These machines use a vacuum to simulate puffing, collecting mainstream smoke separately as particulate matter and gases, followed by further analytical techniques to detect and quantify specific chemicals in those fractions. A study measuring puffing parameters (such as puff volume, duration and inter-puff interval) for little cigars and cigarillos showed that these parameters could be accurately replicated using adapted protocols in cigarette smoking machines.66 Roll-your-own cigarettes

Roll-your-own cigarettes are made from dried tobacco leaves rolled in papers by the user and may be smoked with or without a filter.68 Roll-your-own cigarettes can be made manually by the user or using a cigarette rolling machine.69 While the construction of factory-made cigarettes is highly standardised, roll-your-own cigarettes vary considerably between users. People who smoke roll-your-own cigarettes are reported to take more puffs and inhale more smoke per cigarette, and inhale for a longer duration during smoking.68 ,70

See Section 12.2.2 for further description of roll-your-own tobacco products and Section 3.27.1 for the health effects.

The emissions from roll-your-own cigarettes (that are machine-made or hand-made) can be analysed using a cigarette smoking machine.71 The amount of nicotine and other chemicals varies over different users, different weights of tobacco in the cigarette, the type of paper used (with differing porosities) and whether a filter is used.71 The rolling machine used to make roll-your-owns uses a tube to make standard sizes. Differences in this tube size affect the amount of tar delivered, as measured by a smoking machine.72 Waterpipes

Waterpipes heat tobacco using burning coals placed above the tobacco, to produce emissions that are passed through water before inhalation (see Section Waterpipe tobacco is heated but not directly set alight, therefore doesn’t undergo combustion. However, waterpipe emissions are a mix of those from the heated tobacco and the burning coals that heat the tobacco, which are undergoing combustion. See Section 12.2.5 for the tobacco used in water pipes and Section 3.27.5 for the health effects of waterpipe use.

Waterpipe use is challenging to study in laboratory experiments due to the great variety of ways in which waterpipes are constructed and used. Puff topography studies of waterpipe users have revealed great variation in puffing parameters, with between 500 to 1000 ml volumes, 2- to 3-second puff durations and 10 to 35 seconds of inter-puff intervals.74 Waterpipe sessions also do not have a defined ending time; rather the users usually finish when they perceive a change in conditions that makes the tobacco taste less favourable.74

An International Organization for Standardization (ISO) standard procedure was developed for measuring emissions from waterpipes in 2019, providing the specifications for an analytical “research grade” waterpipe puffing machine.75 This standard uses electrical heating of waterpipe tobacco at 280°C (appropriate for pyrolysis but not combustion, see Sections and using a standardised waterpipe configuration of a laboratory glass bottle, a fixed volume of water in the water bowl, and plastic tubes that emulate the waterpipes used in the community. Vacuum suction is applied to simulate inhalation from which gases and particles are recovered from mainstream smoke for further analysis. The standard puffing parameters are 175 puffs of 530 ml, taken every 20 seconds, with a puff duration of 2.6 seconds, similar to the “Beirut method”.76 ,77 However, charcoal is not used in this standard protocol, with emissions coming solely from the heated tobacco. Standard operating procedures for measuring the emissions of specific chemicals in cigarettes have been determined by the WHO’s Tobacco Laboratory Network (see Section 12.5.3). These are suitable, once modified, for determining the amounts of chemicals such as nicotine, carbon monoxide and nitrosamines in emissions from waterpipe tobacco. But these standard operating procedures are not suitable for measuring charcoal constituents.74

Considerable research has been conducted prior to the generation of the ISO waterpipe standard procedure, which used a variety of waterpipe configurations and puffing topology parameters.74 Increasing the puff volume results in increased consumption of tobacco, perhaps because the increased airflow leads to a higher temperature in the tobacco.74 ,78 A higher temperature, with lower humectants in the tobacco, delivered an increased amount of toxic carbonyls produced in the smoke, such as acetaldehyde, formaldehyde, acetone and acrolein.79

Exposure of the waterpipe user to emissions differs from the research grade experiments in a number of ways. The toxicants produced by waterpipes are different if charcoal is used to heat the tobacco, as emissions from the charcoal constitute part of the total emissions. Users are also exposed to secondhand smoke from sidestream smoke from the charcoal and tobacco, plus smoke exhaled by other users.74 Bidis

Bidis are small, thin, hand-rolled cigarettes that are popular in Asia. They consist of tobacco flakes rolled in a piece of dried tendu or temburni leaf (from plants native to Asia).80 Bidis are mass-produced in a labour-intensive process which is more variable than the highly automated production of factory-made cigarettes. To make bidis, the tobacco is hand-rolled in a prepared leaf and secured with a thread. See Section 12.2.7 for more information about the tobacco in bidis and 3.27.7 for the health effects of smoking bidis.

Bidis are a similar size to cigarette, and therefore their emissions can be analysed using smoking machines similar to those for cigarettes. However, the variability in bidi construction makes them more challenging to analyse. Bidis are also used differently than cigarettes, with a higher puff intensity and shorter inter-puff duration. The low combustibility and nonporous nature of the leaf wrappers means that stronger puffs, increased puff volume and increased puff frequency are needed to light a bidi and keep it burning.81 Custom-made smoking machines have been used to analyse the chemicals present in the mainstream smoke from bidis.81 ,82 Harmful and carcinogenic chemicals in bidi emissions were detected using a puff duration of six seconds and a six second inter-puff interval.81

The International Organization for Standardization have developed a standard procedure for machine-smoking of bidis.83 Kreteks

Kreteks are Indonesian cigarettes containing cloves, clove oil and tobacco. Traditional kreteks are hand-rolled in a corn husk wrapper. Contemporary kreteks are mass produced, paper-wrapped and rolled by machine, however hand-rolled and corn-husk wrapped kreteks are also available.84 The tobacco in kreteks is described in Section 12.2.6 and health effects of smoking kreteks in Section 3.27.6.

One study of kretek emissions used a cigarette smoking machine to generate the smoke using a two second puff duration, 35 ml puff volume and one puff every 60 seconds, comparable to conditions for smoking cigarettes. Nicotine, tar and carbon monoxide were detected in the kretek emissions.85 Smokeless tobacco

Smokeless tobacco refers to tobacco products that are used orally or nasally. It is not heated during use, so does not usually contain many of the toxic chemicals produced during pyrolysis and combustion found in tobacco products that are heated or burned. However, some types of smokeless tobacco are fire-cured, roasted or partially burnt during their manufacture, and therefore likely to contain toxic chemicals produced during pyrolysis.23 ,86

Smokeless tobacco produces emissions, which are all the chemicals released when it is used, affecting the mouth or nose of the user. Some of the chemicals from smokeless tobacco, such as nicotine, move across the cellular membranes into the cells lining the mouth or nose, and from there make their way into the blood stream where they circulate around the body. Many of the studies of chemicals in smokeless tobacco use methods that measure these chemicals in the unused product,87-89 rather than emulating the processes of chewing or sniffing the products.

In 2017 the WHO Study Group on Tobacco Product Regulation recommended that the Tobacco Laboratory Network work to adapt the current standard operating procedures for detection of nicotine, tobacco-specific nitrosamines, benzo[a]pyrene and other toxicants for analysis smokeless tobacco.74 The Tobacco Laboratory Network have released two new standard operating procedures for the determination of the pH and moisture content of smokeless tobacco.90 ,91

12.5.6 Biomarkers of tobacco smoke exposure and damage

The purpose of measuring chemicals in tobacco emissions is to allow regulators to gain insight into the relative harm done to human health from exposure to tobacco smoke from different sources. However, as discussed above, tobacco product emissions data are a poor predictor of human exposure and risks. A more accurate approach for measuring the risk of smoke to human health is to measure how much of specific chemicals from tobacco smoke enters the human body. This can be done by detecting biomarkers of exposure. Detection of chemicals from smoke and their metabolites in the body

A biomarker, in the case of tobacco use, is a substance in the human body that can be measured in a test that will indicate exposure to specific chemicals from tobacco smoke. Biomarkers of tobacco exposure can be detected in people’s breath, saliva, urine, blood and other samples. Biomarkers may also indicate the dose of the chemical, the biological activity, damage—or the potential for damage—to human systems, and the risk of disease.92

Most of the chemicals from smoke are immediately changed by chemical reactions once they enter human cells or the blood stream. This process is called metabolism, and the products from metabolism are referred to as metabolites. Directly measuring the levels of the chemicals from tobacco emissions in the body to estimate exposure will therefore be inaccurate; detection of the metabolites of these chemicals will be necessary. Metabolism of chemicals from smoke can produce more than one product, and the amounts of different metabolites might vary from person to person, and over time. In many cases, it is the metabolic products of the chemicals from smoke that actually do harm to human health. The harmful metabolites are often intermediates in a multi-step metabolism process. As metabolism reactions continue, the intermediates are converted into less harmful forms (detoxification). Considerable research effort was necessary, and is ongoing, to validate the use of biomarkers; i.e. to establish biomarker tests that provide a valid estimate of exposure from tobacco smoke, and of the risk of damage and health effects where possible. Biomarker detection in different regions of the body

The majority of exposure to tobacco smoke occurs through the airways. Chemicals from smoke may also enter the body through other routes, such as via the skin, coming from surfaces contaminated with thirdhand smoke (see Section 4.3).93 People inhaling smoke will be exposed to its chemical content via their lips, nose, mouths, throat, larynx (voice box) and lungs. Once in the alveoli of the lungs, the chemicals from smoke can directly enter lung cells and move into the blood stream. In the blood, these chemicals circulate to all parts of the body, and can exit the blood stream into various organs and regions. This can happen very rapidly. For instance, the time it takes for nicotine from a puff of a cigarette to reach the brain is 10 to 20 seconds.94 However, not every chemical from tobacco smoke gains entry into the lung cells, blood stream or the rest of the body, and the different regions of the body may be exposed to different amounts of these chemicals.

Research on biomarkers has established tests for many different chemicals from tobacco smoke. From the breath of smokers, carbon monoxide and nitric oxide can be directly measured, as well as volatile organic compounds such as benzene.95 Metabolites of nicotine and cyanide can be detected in the saliva of smokers.96 ,97 Urinary metabolites are often useful biomarkers of the uptake of chemicals into the body and may also provide information on metabolic activation, detoxification and dose.92 Urinary cotinine is a useful biomarker of nicotine uptake.92 One study has shown that urinary cotinine (a biomarker of nicotine), NNAL (a biomarker of the nitrosamine NNK) and 1-HOP (a biomarker of polycyclic aromatic hydrocarbons) increase with increasing number of cigarettes smoked per day.98 Biomarkers of chemicals from smoke have also been found in the cervical mucous, pancreatic fluid, placenta and seminal fluid from smokers.92 Toxic metals such as cadmium can be found in the blood of smokers, without the need for biomarkers.46 Biomarkers for detection of damage and disease risk

Biomarkers that signal damage to cells as well as the risk of disease are particularly useful. For cancer caused by smoking, common biomarkers are carcinogens bound to specific sections of DNA, called DNA adducts (see Section 3.3.2). DNA adducts have the potential to result in genetic mutations that can contribution to carcinogenesis—the formation of cancer. Many studies have detected higher levels of DNA adducts in the tissues of smokers compared to non-smokers.92 Some human and animal data support an association of increased levels of DNA adducts with the formation of cancer, with research into this area rapidly evolving.92 Biomarkers for the detection of specific chemicals from smoke

A very useful biomarker for nicotine is the detection of Total Nicotine Equivalents. This is the sum of nicotine, cotinine, 3′-hydroxycotinine and other metabolites of nicotine. When measured together in the urine, these constitute 73–96% of the nicotine dose received by a person exposed to tobacco.23 ,99 ,100 Urinary metabolites of nicotine have been shown to be associated with lung cancer in a prospective study.101

Tobacco-specific nitrosamines are potent carcinogens. Their metabolites make useful biomarkers of tobacco exposure as they are only found in tobacco in appreciable levels. They can be used to estimate dose and their exposure is strongly associated with harm.92 A measure called Total NNAL is the sum of levels of a range of NNAL metabolites and is a useful urinary or blood biomarker for NNK.23 ,100 ,102 Total NNAL in serum from the blood or in urine has been shown to be associated with lung cancer in prospective studies.103 ,104

Polycyclic Aromatic Hydrocarbons (PAHs) are a range of carcinogenic compounds found in tobacco smoke, as well as fossil fuels. They are produced when these carbon compounds are burned as well as during the cooking of fatty meats at high temperatures. The metabolite 1-HOP is a biomarker of pyrene, part of a mix of PAHs, and a widely accepted biomarker of PAH exposure.23 ,100 ,102

A panel of useful biomarkers proposed by researchers in 2010100 includes those described above and others such as urinary biomarkers MHBMA (for 1,3-butadiene), SPMA (for benzene) and HPMA (for acrolein), blood biomarkers cyanoethylvaline (for acrylonitrile) and carbamoylethylvaline (for acrylamide) and exhaled carbon monoxide as a direct test for exposure to this gas. These authors also recommend the use of DNA adduct biomarkers for potential damage in white blood cells by exposure to formaldehyde and acetaldehyde.100

The levels of many of the biomarkers described above are quite rapidly reduced when people stop smoking.100 Advantages and disadvantages of biomarkers

There are many advantages to using biomarkers to estimate human exposure to tobacco smoke. Biomarkers should capture overall exposure to tobacco smoke, not just exposure to smoke from the products used by the individual. People who smoke may also be subject to secondhand smoke from other smokers. They may not accurately report the number of cigarettes or the amount of tobacco they regularly consume. Smokers may use more than one type of tobacco product, or smoke different types of products in different ways, such as smoking lower nicotine cigarettes more intensely. They may also be exposed to some of the chemicals in tobacco by chewing tobacco or heated tobacco products. Using a biomarker can estimate the total exposure, taking into account all these variations. Furthermore, there are individual differences in the metabolism of toxic and carcinogenic chemicals from smoke. The enzymes that convert these chemicals to more damaging forms differ in their potency in different people (see Section 3.24.1). This leads to different levels of biologically harmful chemicals. By measuring levels of the harmful chemicals, these differences can be taken into account, in an effort to more accurately predict harm. Biomarkers may also be used to test the dose of chemicals that reach specific parts of the body. Dose of exposure to a toxic compound is related to its toxicity.

There are disadvantages of using biomarkers to measure exposure from tobacco smoke. Many biomarkers are not specific to tobacco smoke—because people are exposed to the same chemicals from other sources. For instance, people are exposed to toxic acrolein from tobacco smoke but also from fatty foods cooked at high temperatures. The metabolite 3-hydroxypropyl mercapturic acid (HPMA) is a useful biomarker for acrolein, but measuring this will also detect acrolein exposure from other sources.105 Experiments measuring such biomarkers need to take into account these other exposures. There are biomarkers that are specific for tobacco, such as the metabolite NNAL, a biomarker of NNK—a tobacco-specific n-nitrosamine.102 Another disadvantage of using biomarkers is that all of the metabolites of a chemical may not be known or readily measurable, making it difficult to define an accurate panel of markers for some tobacco chemicals. Using a biomarker of potential harm, such as a DNA adduct, is one way of addressing this issue.100 A DNA adduct is a carcinogen bound to specific sections of DNA (see Section 3.3.2). DNA adducts may cause genetic mutations that can contribute to the formation of cancer.92 Some human and animal data support an association of increased levels of DNA adducts with the formation of cancer.92 Protein adducts can also be measured, where a metabolite of a smoke chemical has bound to a protein in a way that has the potential to cause damage. Measuring the level of DNA and protein adducts, rather than the level of metabolites, may be closer to estimating the damage done by smoking than the amount of specific metabolites of chemicals from smoke.92


Relevant news and research

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