Cigarette smoke inhalation and blood dose markers in beagle dogs

The time course of three tobacco-related blood dose markers was determined in beagle dogs during and after two-, four-, and six-cigarette acute smoke exposures. Venous blood samples were taken from a jugular catheter at selected intervals and analyzed for nicotine, cotinine, and carboxyhemoglobin (COHb). Smoke was generated from reference cigarettes (1.91 mg of nicotine) by a machine and delivered to the trachea of tracheostomized dogs. Cigarettes were chain-smoked in pairs with a 5-min break between pairs when four or six cigarettes were smoked. Blood nicotine, cotinine, and COHb levels all rose during smoke inhalation. Peak values of COHb were dosimetrically related to the number of cigarettes, while peak nicotine and cotinine values were not related consistently. The exposure protocol used in this study produced blood levels of all three measured parameters which were in the range of published values for human smokers, so that it is possible to approximate acute human cigarette smoke inhalation in the beagle dog model.     

Population characteristics and cigarette yield as determinants of smoke exposure

Relationships of population characteristics, smoking history, and cigarette yield with smoke exposure as measured by peripheral blood concentrations of thiocyanate, carboxyhemoglobin, nicotine and cotinine were sought in 170 male smokers. This population of smokers had significant elevations of serum thiocyanate, blood carboxyhemoglobin and plasma nicotine and cotinine concentrations as compared with an equal number of age- and sex-matched nonsmokers and these concentrations correlated significantly with past 24-hour cigarette consumption. Although the nicotine yield of the cigarette correlated significantly with plasma cotinine and marginally with plasma nicotine, the reduction in plasma nicotine and cotinine was not proportionate to the reduced yield of the cigarettes, suggesting that smokers partially compensate for the lower yields of their cigarettes. Blood levels of carboxyhemoglobin, nicotine and cotinine were also significantly associated with the weight of the subjects, presumably due to the relationship between weight and the volume of distribution. Univariate and multiple regression analyses provided evidence that coffee and alcohol consumption and years smoked also may be important determinants of smoke exposure.     

Effects of cigarette nicotine content on smoking behavior of baboons

Human cigarette smokers modify the way in which they smoke cigarettes of differing nicotine content, apparently to maintain nicotine exposure at a preferred level. The effects of changing from moderate to high or low nicotine content cigarettes were examined in 11 baboons (superspecies Papio cynocephalus) trained to smoke cigarettes for water rewards. Relative to the moderate nicotine content cigarette, the animals took significantly (p < .05) more puffs on the high nicotine content cigarette, and the puffs on the high nicotine cigarette were significantly larger in volume. The animals made the same number of puffs, relative to the moderate nicotine content cigarette, on the low nicotine content cigarette, but the volume of the puffs was significantly smaller. The cotinine output in urine varied significantly and was directly related to cigarette nicotine content; cotinine is the primary metabolite of nicotine. Baboons, like people, prefer high nicotine content cigarettes. Nonhuman primates also regulate nicotine exposure by modification of their puffing behavior. These results indicate that the nonhuman primate also can be used as a model for the investigation of the behavioral aspects of cigarette smoking.     

Pharmacokinetics of nicotine in rats after multiple-cigarette smoke exposure

The pharmacokinetics of nicotine and its major metabolites was evaluated in male rats after multiple-cigarette smoke exposure. A smoke-exposure apparatus was used to deliver cigarette smoke to the exposure chamber. The rats were exposed to smoke from a single cigarette every 8 hr for 14 days and to the smoke of a cigarette spiked with radiolabeled nicotine on the 15th day. Blood and urine samples were collected at timed intervals during the 10-min smoke-exposure period of the last cigarette and up to 48 hr thereafter. Nicotine, cotinine, and other polar metabolites were separated by thin-layer chromatography and quantified by liquid scintillation counting. The data were analyzed by computer fitting, and the derived pharmacokinetic parameters were compared to those observed after a single iv injection of nicotine and after a single-cigarette smoke exposure. The results indicated that the amount of nicotine absorbed from multiple-cigarette smoke was approximately 10-fold greater than that absorbed from a single cigarette. Also, unlike the single-cigarette smoke exposure experiment, nicotine plasma levels did not decay monotonically but increased after the 5th hr, and high plasma concentrations persisted for 30 hr. The rate and extent of the formation of cotinine, the major metabolite of nicotine, were decreased as compared with their values following a single-cigarette smoke exposure. It was concluded that nicotine or a constituent of tobacco smoke inhibits the formation of cotinine and may affect the biotransformation of other metabolites. Urinary excretion tended to support the conclusions that the pharmacokinetic parameters of nicotine and its metabolites were altered upon multiple as compared to single dose exposure.     

Hydroxyethylvaline adduct formation in haemoglobin as a biological monitor of cigarette smoke intake

The ethylene oxide adduct formed on the N-terminal valine in haemoglobin was investigated as a biological monitor of tobacco smoke intake. The modified method developed for the determination of the hydroxyethylvaline adduct (HOEtVal) involved reaction of globin with pentafluorophenyl isothiocyanate, extraction of the HOEtVal thiohydantoin product, derivatization of this by trimethylsilylation and quantitation by capillary gas chromatography with selective ion monitoring mass spectrometry using a tetradeuterated internal standard. The method was applied to globin samples from 26 habitual cigarette smokers and 24 non-smokers. There was a significant correlation between cigarette smoke intake as measured by the average number of cigarettes smoked per day and HOEtVal levels (r=0.537, p<0.01). Background levels were found in non-smokers (mean 49.9 pmol/g Hb, range 22–106 pmol/g Hb). Smoking increased these levels by 71 pmol/g Hb/ 10 cigarettes per day.Cotinine levels in plasma of the smokers were determined by GC-NPD using 2-methyl-4-nitroaniline as internal standard. For non-smokers cotinine was determined by GC-MS selective ion monitoring using d₃-methylcotinine as internal standard. There was no correlation between number of cigarettes smoked per day and cotinine levels (r=0.297, p>0.05) although cotinine was correlated with HOEtVal (r=0.43, p<0.01).The HOEtVal adduct levels thus appear to be a suitable biomonitor for exposure to hydroxyethylating agents in cigarette smoke, reflecting an integrated dose over the erythrocyte lifetime. This is in contrast to plasma cotinine determinations which reflect only the previous day's exposure to nicotine in smoke.     

The effects of a restricted feeding regimen on cigarette smoking in humans.

The effects of 3 days of restricted feeding (800 kcal/day) on cigarette consumption, smoke exposure, and mood were studied in five male research volunteers. A within-subjects design was used in which subjects were exposed in an inpatient research unit to either a nutritionally-balanced diet containing 800 kcal (RESTRICTED DIET) or 3,000 kcal (NORMAL DIET) per day for 3 consecutive days. At least 2 weeks separated diet conditions. Dependent measures included number of cigarettes smoked per day in each diet condition, biological exposure levels (carbon monoxide and plasma cotinine levels), and mood. Number of cigarettes smoked per day did not differ significantly across diet conditions. Biological exposure to carbon monoxide and to cotinine, a metabolite of nicotine indicative of chronic nicotine exposure, also did not differ significantly between conditions. Fatigue scores from the Profile of Mood States were significantly elevated in the RESTRICTED DIET condition. Not surprisingly, subjects in this condition also reported feeling more hungry throughout the day than in the NORMAL DIET condition. From our study results, we conclude that a short period of "dieting," and the resulting hunger elicited from such a diet, do not increase cigarette consumption or smoke exposure in humans.     

Passive and active exposure to cigarette smoke in a smoking experiment

Six volunteer female habitual smokers were exposed during a 2-wk experimental period to cigarette smoke, both actively and passively, in an exposure chamber (volume 10 m3, average air exchange rate 6.8 times/h), where the ambient carbon monoxide, particle, and aldehyde concentrations were monitored. Three of the six subjects were smoking at the time, 2 cigarettes (filtered, self-burning low tar brand) per person per hour, 30 cigarettes altogether during each of the 5-h experimental days in the chamber. Samples of blood and urine were taken from each subject after 3 nonsmoking days and after each day of active or passive smoking. Among the parameters tested, blood carboxyhemoglobin, plasma cotinine, and urinary mutagenicity were higher in samples taken after active smoking than after nonsmoking periods. Although the exposure conditions were similar for all subjects, the parameters measured showed quite high interindividual variation. Thioethers and thiocyanates were not significantly elevated in the active smoking samples; neither were there any differences during this short experimental period in the sister chromatid exchange frequencies. The only parameters showing an increasing trend after passive exposure, as compared with nonsmoking samples, were urinary mutagenicity and plasma cotinine, the main metabolite of nicotine.     

Cigarette smoking as a risk for cardiovascular disease III: Biochemical effects with higher nicotine yield cigarettes

Subjects who smoked a medium range nicotine yield cigarette were given a higher nicotine yield cigarette (an increase of 0.34 mg nicotine) to smoke ad libitum for two weeks. Plasma nicotine, cotinine, thiocyanate and blood carboxyhemoglobin levels were determined as well as various physiological parameters including heart rate and blood pressure. Increases in plasma nicotine were most directly correlated to heart rate when smokers were first challenged with a higher nicotine yield cigarette (r = 0.85); less directly correlated after a two-week acclimatization period (r = 0.42) and poorly related to their customary product (r = 0.23). Interestingly, it was noted that subjects did not compensate for higher nicotine yield by smoking fewer cigarettes per day when incremental nicotine changes were realistic. They did, however, show higher plasma nicotine, thiocyanate and an upward trend in plasma cotinine with the stronger cigarettes. These increases in cigarette constituents present in plasma, coupled with increasing correlation of heart rate and nicotine uptake, lead us to suggest that uptitration of smokers might cause them to establish new baseline levels. These findings have important health implications in light of recent suggestions to increase the nicotine yet decrease the tar of cigarettes in an attempt to overcome smoker compensation phenomena observed with low yield products.     

Long-term effects of switching to cigarettes with lower tar and nicotine yields

On switching to cigarettes with lower tar and nicotine yields, most individuals smoke more intensively, but it is not clear if this effect persists over a long period. Smoking behaviour was monitored in 10 male and 18 female volunteers at five monthly visits, smoking commercially available cigarettes (tar yield>10 mg), then for six more visits at 6-week intervals after switching (mean reduction of 5.9 mg tar and 0.45 mg nicotine). Puffing behaviour was monitored with a flow sensing holder, and measurements were made before and after smoking of plasma cotinine, carboxyhaemglobin and alveolar carbon monoxide. After switching, cotinine levels only fell 40% of that predicted from the fall in nicotine yields, and there were no systematic trends for the rest of the study. Puff volumes rose (reflecting perhaps the reduced draw resistance of the lower yield cigarettes), and remained higher thereafter. The number of puffs per cigarette appeared to rise on switching, but then decreased again. In conclusion, most effects of switching to lower yield cigarettes appeared to persist for at least 36 weeks, suggesting that the strategy of reducing exposure to cigarette smoke by lowering tar and nicotine yields may be of limited value.     

Analytical cigarette yields as predictors of smoke bioavailability

The smoke intake of 865 undisturbed smokers of over 10 cigarettes per day was measured using plasma nicotine and cotinine, and expired carbon monoxide (CO) as markers. While nicotine yields, according to Federal Trade Commission (FTC) analytical standards, varied 16-fold from 0.1 to 1.6 mg/cigarette, the corresponding plasma nicotine values varied from around 25 to 45 ng/ml, and estimated mean nicotine intake of smokers varied from around 0.75 to 1.25 mg/cigarette. Expired CO and plasma cotinine values also varied in similar proportion, but mean daily cigarette consumption was independent of the FTC nicotine yield of the cigarettes smoked. The results indicate that pharmacodynamic satiation causes behavioral regulation, and that smokers of very high yield brands compensate downward, and vice versa. The ratio of tar yield to nicotine yield usually increases with increasing tar yield; therefore tar intake is likely to increase at higher tar yields, even though the increment of nicotine intake is small. It follows that FTC analytical determinations are poor predictors of relative intake of nicotine, CO, or tar, while rankings based on mean tar-to-nicotine ratio of a brand's smoke could be more meaningful. Moreover, the considerable variation of individual smoking behavior suggests that precise numerical rankings of cigarettes are not justified. An analogic ranking of cigarettes into a few broad classes would better reflect the realities and expectations of average consumers.     

Cigarette smoking as a risk for cardiovascular disease V: Biochemical parameters with increased and decreased nicotine content cigarettes

Cigarette smokers were assessed for customary smoking behavior and then were assigned a cigarette which was 0.4 mg higher or lower in nicotine and after 4 weeks, were returned to their customary brand. Biochemical indices of smoking behavior including blood carboxyhemoglobin (COHb), plasma nicotine, cotinine and thiocyanate (-SCN) were measured every 2 weeks. When nicotine availability was increased, smokers received an increased nicotine bolus per puff as determined by plasma nicotine and did not alter smoking topography or cigarettes per day. Over the 4 weeks, plasma cotinine increased without corresponding increases in COHb and -SCN. The return to standard brand resulted in declining cotinine levels but increasing COHb and -SCN, suggesting altered inhalation patterns. In smokers switched to a low yield cigarette, there was a decrease in the nicotine obtained per cigarette followed by a steady rise in plasma cotinine, -SCN and blood COHb over the 4-week period. A positive correlation was observed between cotinine and the gas phase constituents during the change to lower yield and back to standard brand cigarettes. These results indicate that cigarette smokers compensate for decreased nicotine yield with concomitant increases in gas phase components. In addition, increased nicotine availability results in an increased body burden of nicotine and “tar,” but not gas phase constituents. The relative risks of cardiovascular disease under these two situations, which increase exposure to nicotine or gas phase components, deserve careful consideration.     

Blood nicotine, smoke exposure and tobacco withdrawal symptoms

The relationship between tobacco withdrawal symptoms and pre- and post-cigarette blood nicotine levels, pre-cigarette cotinine levels, change in nicotine level from pre- to post-cigarette, half-life for nicotine, and total smoke exposure was examined in 20 smokers. Subjects' reports of craving for cigarettes were significantly related to blood nicotine/cotinine levels and change in nicotine level from pre- to post-cigarette; questionnaire measures of confusion and number of awakenings during sleep was related to half-life for nicotine; and number of awakenings during sleep was related to behavioral measures of total smoke exposure. These results suggests some symptoms of tobacco withdrawal are related to nicotine deprivation while others are not.     

Good relationship between saliva cotinine kinetics and plasma cotinine kinetics after smoking one cigarette

This study investigated the relationship between plasma and saliva cotinine kinetics after smoking one cigarette and the relationship between cotinine kinetics and estimated nicotine intake, which was calculated as mouth level exposure (MLE) of nicotine, from smoking two test cigarettes with different nicotine yields. This study was conducted in 16 healthy adult Japanese smokers, who did not have null nor reduced-activity alleles of CYP2A6, with a quasi-randomized crossover design of smoking a low-tar cigarette or a high-tar cigarette. Saliva cotinine showed similar concentration profiles to plasma cotinine, and all of the calculated pharmacokinetic parameters of cotinine showed the same values in plasma and saliva. The C_max and AUC of cotinine showed almost the same dose-responsiveness to the estimated MLE of nicotine between plasma and saliva, but the t_max and t_1/2 of cotinine were not affected by the estimated MLE of nicotine in either plasma or saliva. The results show that saliva cotinine kinetics reflects plasma cotinine kinetics, and measurement of saliva cotinine concentration gives the same information as plasma cotinine on the nicotine intake. Thus, saliva cotinine would be a good and less-invasive exposure marker of cigarette smoke, reflecting the plasma cotinine concentration and kinetics.     

Urinary cotinine levels of electronic cigarette (e-cigarette) users

The popularity of electronic cigarettes (e-cigarettes) is rapidly increasing in many countries. These devices are designed to imitate regular cigarettes, delivering nicotine via inhalation without combusting tobacco but currently, there is a lack of scientific evidence on the presence or absence of nicotine exposure. Such research relies on evidence from e-cigarette users urine samples. In this study, we aimed to determine the levels and compare the amount of nicotine to which e-cigarette users, cigarette smokers and passive smokers are exposed. Therefore, urine samples were collected from e-cigarette users, cigarette smokers, passive smokers, and healthy nonsmokers. The urinary cotinine levels of the subjects were determined using gas chromatography–mass spectrometry. The mean (±SD) urinary cotinine levels were determined as 1755?±?1848?ng/g creatinine for 32 e-cigarette users, 1720?±?1335?ng/g creatinine for 33 cigarette smokers and 81.42?±?97.90?ng/g creatinine for 33 passive smokers. A significant difference has been found between cotinine levels of e-cigarette users and passive smokers (p?<?0.05). There were no statistically significant differences between e-cigarette users and cigarette smokers (p?>?0.05). This is a seminal study to demonstrate the e-cigarette users are exposed to nicotine as much as cigarette smokers.     

Influences of exposure to cigarette smoke on concentration of nicorandil in plasma of rats.

Influences of acute exposure to cigarette smoke on plasma concentrations of nicorandil administered orally and parenterally were investigated in rats by HPLC. The animals were exposed to tobacco smoke of two kinds of cigarettes using a smoking machine (i.e., the cigarette smoke contained either low or high nicotine and tar). The plasma concentration of nicorandil administered orally at a dose of 10 mg/kg had a lower absorption phase in two cigarette smoke-exposed groups, particularly in the high nicotine and tar-containing cigarette smoke-exposed group, compared with the nonsmoking control group. The AUC and MRT values in a high nicotine and tar-containing cigarette smoke-exposed group were lower and higher, respectively, than in the nonsmoking control group. However, there was no marked difference in nicorandil plasma concentrations between the cigarette smoke-exposed group and the nonsmoking control group when nicorandil was administered ip or iv at a dose of 5 mg/kg. These results suggest that cigarette smoke exposure causes the suppression or delay of absorption of nicorandil from the gastrointestinal tract.     

Development and Characterization of an Elisa for Cotinine in Biological Fluids

Abstract

Cotinine is a major metabolite of nicotine and serves as an important biomarker of tobacco smoke exposure. To monitor exposure to tobacco smoke or nicotine, a sensitive enzyme-linked immunosorbent assay (ELISA) for cotinine was developed. The test had an 1–50 of between 0.5 and 1.0 ng/ml for cotinine and about 500-fold less affinity for nicotine. Few matrix effects were not detectable in human saliva, although relatively small matrix effects (1–50 for cotinine, 2.8 ng/ml) were observed in human serum and urine. The test accurately measured the levels of cotinine in NI5T standards in freeze-dried human urine derivative material (r = .9999) indicating its reliability for measurement of cotinine. The test readily detected low levels (5–500 nglml) of cotinine in human saliva and serum samples. Also, the levels of cotinine in plasma and urine samples from smoke-exposed mice and rats could be rapidly analyzed for cotinine. This ELISA is therefore a sensitive and accurate test for the determination of cotinine in plasma, serum, saliva, and urine samples from humans and animals, and can be successfully used for monitoring and quantifying exposure to tobacco smoke or nicotine.     

Hair analysis--a biological marker for passive smoking in pregnancy and childhood.

Passive smoking has been shown to adversely affect the health of infants and children. We used hair analysis for nicotine and its metabolite cotinine as a biological marker for exposure to smoking in these two groups. Using radioimmunoassay we measured maternal and fetal hair concentrations of nicotine and cotinine in the mother-infant pairs belonging to three different groups based on the mother's smoking habits. The three groups were: active smokers, passive smokers and nonsmokers. There was a significant correlation between maternal and neonatal hair concentration for both, nicotine and cotinine. Mothers and infants in the smoking groups, both active and passive, had significantly higher hair concentrations of both, nicotine and cotinine than in the control, nonsmoking group. In an older cohort, we compared two groups: 78 asthmatic children were compared to 86 healthy children exposed to similar degrees of passive smoking. By using objective, biological markers, our study aimed at verifying whether asthmatic children are different from nonasthmatic children in the way their bodies handle nicotine. Our results show, that, despite the fact that parents of asthmatic children tend to smoke a lower number of cigarettes per day, their children had an average twofold higher concentrations of cotinine in their hair then the control, nonasthmatic children. These studies document the importance of hair analysis as a tool for measuring exposure to cigarette smoke.     

The effect of menthol containing cigarettes on adult smokers’ exposure to nicotine and carbon monoxide

There is limited information comparing biomarkers of exposure (BOE) to cigarette smoke in menthol (MS) and non-menthol cigarette smokers (NMS). Objective: To compare BOE to nicotine and carbon monoxide in MS and NMS. Methods: Cross-sectional, observational, ambulatory, multi-centre study in 3341 adult cigarette smokers. Nicotine equivalents (NE) in 24 h urine, NE/cigarette, COHb and serum cotinine were measured. Statistical analyses included analysis of variance and Wilcoxon test. Results: Analyses of variance revealed no statistically significant effects of mentholated cigarettes on NE/24 h, COHb, serum cotinine and NE/cigarette. On average MS smoked 15.0 and NMS 16.8 cigarettes/day. The unadjusted mean differences were as follows: MS had lower NE/24 h (5.4%) and COHb (3.2%), higher serum cotinine (3.0%) and NE/cigarette (5.7%) than NMS. African-Americans MS smoked 40% fewer cigarettes, showed lower NE/24 h (24%) and COHb (10%) and higher NE/cig (29%) and serum cotinine (8%) levels than their White counterparts. Conclusions: Smoking mentholated cigarettes does not increase daily exposure to smoke constituents as measured by NE and COHb. These findings are consistent with the majority of epidemiological studies indicating no difference in smoking related risks between MS and NMS.     

Recovery cycle of the Hoffmann reflex of tobacco smokers and nonsmokers: relationship to plasma nicotine and cotinine levels

Ten normal adult tobacco smokers and 10 non-smoking volunteers 20–31 years of age were the subjects of this study. The tobacco smokers all had a history of daily tobacco smoking. They were asked to stop smoking for 12 hours prior to testing. The Hoffmann (H) reflex and its recovery cycle were measured on different days before and just after smoking one nonfiltered 0 mg, low (0.27 mg), or high (2.16 mg) nicotine containing cigarette in a randomized order. Blood samples were drawn immediately after the H reflex recordings in the tobacco smokers. The blood samples were centrifuged, the plasma removed, frozen, and later assayed for nicotine and cotinine levels.Nonsmokers compared to tobacco smokers before smoking only had a tendency for enhanced amplitude of the recovery cycle. After smoking the nicotine containing cigarettes, the tobacco smokers had a depression of the amplitude of the H reflex recovery cycle. The amplitude of the H reflex recovery cycle at 160 ms was reduced. This decreased significantly with increasing plasma nicotine and cotinine concentrations. Individual differences were marked.The data obtained are consistent with evidence in animals that nicotine and tobacco smoke stimulate Renshaw inhibitory neurons in the spinal cord, either directly or indirectly which leads to a skeletal muscle relaxant effect.     

Ultralow-yield cigarettes and type of ventilation: The role of ventilation blocking

Habitual smokers of perforation-ventilated cigarettes and of channel-ventilated cigarettes (18 male and 18 female subjects each; nicotine yield 0.1–0.3 mg, 0.2 mg, respectively) were compared with respect to different smoke exposure indicators and puffing behavior. The role of ventilation blocking was assessed by comparing normal lip contact smoking with smoking through a cigarette holder. The presmoking concentrations (plasma nicotine, cotinine, respiratory CO) were higher for channel-filter than for perforation-ventilated cigarettes, as were the pre- to postsmoking boosts (nicotine, CO) with normal lip smoking. Holder smoking resulted in lower boosts than lip smoking for the channel filter cigarettes, although the puffing behavior was considerably intensified. The boosts for perforation-ventilated cigarettes remained unchanged and were reached with only moderately intensified puffing behavior. The results indicate the importance of ventilation blocking in everyday lip smoking for channel-filter cigarettes, but not for conventional, perforated cigarettes.