Urine Of Smokers Research Articles

Nicotine and cotinine in adults' urine: The German Environmental Survey 1998.

In 1998, the German Environmental Survey (GerES III) recruited approximately 5000 adults between the ages of 18 and 69 years. The study population for these analyses consisted of 1580 smokers (34% of the total population) and 3126 nonsmokers. Nicotine and cotinine concentrations in urine were determined by HPLC methods with UV-detection and corrected for creatinine. Nicotine and cotinine concentrations differed between smokers and nonsmokers by factors of 10-100. The multiple linear regression models used for the analyses of nicotine detection in the urine of smokers explained 43.2% and 42.3% of the total volume-specific and creatinine-specific variances, respectively. Cigarette smoking was the major factor responsible for 41% of the total variance. The explained variances of the cotinine results were larger, 51.0% and 49.3% of the total variance were volume-specific and creatinine-specific, respectively. More than 20% of nonsmokers in GerES III were exposed to environmental tobacco smoke at home, at work or in other places. The logistic regression analysis approach used for the group of nonsmokers showed the greatest effects for those exposed to tobacco smoke at home (adjusted OR varied between 4 and 6). These results were seen for nicotine as well as for cotinine excretion. Exposure to tobacco smoke in the workplace doubled the risk for the detection of nicotine and cotinine in urine. When other risk factors such as age, sex, social status, community size, season of urine collection, and the consumption of food containing nicotine such as potatoes, cabbage, tea were included, the effect estimates for tobacco smoke exposure remained unchanged. A new federal bill to diminish environmental tobacco smoke (ETS) exposure in the workplace was recently passed in Germany, but protection of nonsmokers from smoking family members at home needs more attention.

N-Glucuronidation of trans-3‘-Hydroxycotinine by Human Liver Microsomes

trans-3'-Hydroxycotinine is the major nicotine metabolite excreted in the urine of smokers and other tobacco or nicotine users. On average, about 30% of the trans-3'-hydroxycotinine in urine is present as a glucuronide conjugate. The O-glucuronide of trans-3'-hydroxycotinine has been isolated from smokers urine and appears to be the major glucuronide conjugate of trans-3'-hydroxycotinine in urine. However, nicotine and cotinine are both glucuronidated at the nitrogen atom of the pyridine ring. We report here that human liver microsomes catalyze both the N-glucuronidation and the O-glucuronidation of trans-3'-hydroxycotinine. The N-glucuronide was purified by HPLC, and its structure was confirmed by NMR. Both N- and O-glucuronidation of trans-3'-hydroxycotinine were detected in 13 of 15 human liver microsome samples. The ratio of N-glucuronidation to O-glucuronidation was between 0.4 and 2.7. One sample only catalyzed N-glucuronidation, and one sample did not catalyze either reaction. The rates of N-glucuronidation varied more than 6-fold from 6 to 38.9 pmol/min/mg protein; rates of O-glucuronidation ranged from 2.8 to 23.4 pmol/min/mg protein. The rate of trans-3'-hydroxycotinine N-glucuronidation by human liver microsomes correlated well with both the rate of nicotine and the rate of cotinine N-glucuronidation. trans-3'-Hydroxycotinine O-glucuronidation correlated with neither nicotine nor cotinine N-glucuronidation. These results suggest that the same enzyme(s) that catalyzes the N-glucuronidation of nicotine and cotinine may also catalyze the N-glucuronidation of trans-3'-hydroxycotinine in the human liver but that a separate enzyme catalyzes trans-3'-hydroxycotinine O-glucuronidation.

Determination of nicotine and its metabolites in urine by solid-phase extraction and sample stacking capillary electrophoresis-mass spectrometry

The combination of capillary electrophoresis (CE) and mass spectrometry (MS) with solid-phase extraction (SPE) has been used for the identification of nicotine and eight of its metabolites in urine. The recovery of cotinine from cotinine-spiked urine, by C18 SPE, was found to be 98%. Smokers urine (200 ml) was preconcentrated 200-fold via SPE prior to analysis. The sample stacking mode of CE, when compared to capillary zone electrophoresis, was shown to improve peak efficiency by 132-fold. The combination of hydrodynamic and electrokinetic injection was studied with sample stacking/CE/MS. The on-column limits of detection (LOD) of nicotine and cotinine, by this technique, were found to be 0.11 and 2.25 μg/ml, respectively. Hence, LODs of nicotine and cotinine in urine after 200-fold preconcentration were 0.55 and 11.25 ng/ml, respectively.

Effect of pH on mutagenicity of urine from smokers and nonsmokers

Cigarette smoking is a major risk factor for bladder cancer. Some chemicals from cigarette smoke e.g. aromatic amines, are metabolized in the liver and excreted in urine as either glucuronide or acetyl conjugates. These metabolites undergo pH mediated activation to reactive nitreneum ions that may lead to DNA adducts, initiating bladder tumorigenesis in some smokers. We hypothesize that changing the pH of smokers urine will impact the mutagenic outcome of these metabolites. Overnight samples were collected from smokers ( n=11) and nonsmokers ( n=11) and stored at −70 °C. Each urine sample was adjusted to neutral (7.0), basic (8.2) and acidic (5.5) pH. Mutagenic activity was assessed with the Ames test utilizing the Salmonella Typhimurium strain YG1024 microsuspension assay. Urine from smokers was significantly mutagenic compared with nonsmokers ( P<0.001). In both the groups, there was no difference between urine adjusted at different pH ( P>0.05). We conclude that changing the pH of smokers urine might not affect their subsequent risk for bladder cancer.

Simultaneous determination of nicotine and eight nicotine metabolites in urine of smokers using liquid chromatography–tandem mass spectrometry

A method based on liquid chromatography tandem mass spectrometry (LC–MSMS) applying atmospheric pressure chemical ionisation (APCI) in the positive ion mode was developed for the direct determination of nicotine, cotinine, trans-3′-hydroxycotinine, their corresponding glucuronide conjugates as well as cotinine- N-oxide, norcotinine, and nicotine- N′-oxide in the urine of smokers. The assay involves filtration of crude urine, fast liquid chromatography on a reversed-phase column and mass-specific detection using MSMS transitions. Deuterium-labeled nicotine, cotinine, and trans-3′-hydroxycotinine were used as internal standards. Glucuronides used as reference material were either chemically (cotinine- N-glucuronide) or enzymatically synthesized (nicotine- N-glucuronide and trans-3′-hydroxycotinine- O-glucuronide). Precision for the major nicotine analytes at levels observable in urine of smokers was better than 10%. Accuracy expressed in recovery rates in urine matrix for nicotine, cotinine, trans-3′-hydroxycotinine, and cotinine- N-glucuronide ranged from 87 to 113%. Quantitative results for the three glucuronides in urine samples of 15 smokers were compared to an indirect method in which the aglycons were determined with gas chromatography and nitrogen-selective detection (GC–NPD) before and after enzymatic splitting of the conjugates. Good agreement was found for cotinine- N-glucuronide (coefficient of variation, CV: 9%) and trans-3′-hydroxycotinine- O-glucuronide (CV: 20%), whereas the accordance between both methods was moderate for nicotine- N-glucuronide (CV: 33%). The described LC–MSMS method allows the simultaneous determination of nicotine and eight of its major metabolites in urine of smokers with good precision and accuracy. Since the method requires a minimum of sample clean-up and a very short time for chromatography (3 min), it is suitable for determining the nicotine dose in large-scale human biomonitoring studies.

Tobacco-smoke exposure indicators and urinary mutagenicity

In this study, the correlation of indicators of external (i.e. mean daily intake of condensate, nicotine, tobacco and tobacco proteins, and daily number of cigarettes smoked) and of internal tobacco-smoke exposure (i.e. urinary 1-pyrenol, nicotine and its metabolites and trans, trans-muconic acid) with urinary mutagenicity, detected on YG1024 Salmonella typhimurium strain with S9, were examined in 118 smokers. An increase in urinary mutagenicity was clearly significantly correlated with each external and internal indicators of exposure to tobacco smoke (correlation coefficient ( r) ranging between 0.22 and 0.54, P<0.01), with a greater extent in the case of indicators of internal dose. In multiple regression analysis, among the indicators of external exposure, daily tobacco intake was the only variable significantly associated with urinary mutagenicity ( t=2.47, P=0.015, with partial contribution to r 2=5.15%). Instead, when all indicators of exposure (external and internal) were considered in the analysis, the influence of urinary 1-pyrenol on urinary mutagenicity was predominant, followed by those of urinary trans, trans-muconic acid and nicotine plus metabolites ( t=4.63, 2.73 and 2.08, P<0.001, P=0.002 and 0.04, with partial contribution to r 2=17.0, 6.66 and 3.96%, respectively), with no influence at all of external tobacco-smoke exposure indicators. In conclusion, our results show that indicators of internal dose are better correlated with formation of mutagens in urine of smokers. Among these, the best indicator was urinary 1-pyrenol and this result designates the combustion processes of tobacco as the determining step for the formation of urinary mutagens. However, as these biomarkers cannot be analysed the amount of daily tobacco intake represent the best valuable index of external (presumptive) exposure to tobacco-smoke genotoxins.

Determination of cadmium and lead in urine of some Nigerian subjects

Forty-one urine samples of smokers and non-smokers from Ile-Ife, Nigeria were analyzed for normal cadmium and lead levels after acid digestion. Results of the preliminary study show that concentrations of cadmium obtained were generally low compared with lead. The mean urinary levels from non-smokers ranged from 0.05 - 0.02 to 0.16 - 0.01 w g ml -1 and from 0.02 - 0.03 to 0.88 - 0.04 w g ml -1 for cadmium and lead respectively, whilst, for smokers, urinary levels varied between 0.07 - 0.01 and 0.23 - 0.02 w g ml -1 and between 0.28 - 0.03 and 1.02 - 0.02 w g ml -1 were obtained for cadmium and lead, respectively. Cd levels in urine of smokers and non-smokers are higher than the recommended normal level of 0.0085 w g ml -1 , while levels of Pb are above the 0.08-0.15 w g ml -1 exposure range. Results of spiking experiments with urine gave high percentage recoveries for both elements (Cd, 87.9 - 0.40%) and (Pb, 90.40 - 0.12%). Blank determination was done for background correction.

Human urinary carcinogen metabolites: biomarkers for investigating tobacco and cancer

Measurement of human urinary carcinogen metabolites is a practical approach for obtaining important information about tobacco and cancer. This review presents currently available methods and evaluates their utility. Carcinogens and their metabolites and related compounds that have been quantified in the urine of smokers or non-smokers exposed to environmental tobacco smoke (ETS) include trans,trans-muconic acid (tt-MA) and S-phenylmercapturic acid (metabolites of benzene), 1- and 2-naphthol, hydroxyphenanthrenes and phenanthrene dihydrodiols, 1-hydroxypyrene (1-HOP), metabolites of benzo[a]pyrene, aromatic amines and heterocyclic aromatic amines, N-nitrosoproline, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol and its glucuronides (NNAL and NNAL-Gluc), 8-oxodeoxyguanosine, thioethers, mercapturic acids, and alkyladenines. Nitrosamines and their metabolites have also been quantified in the urine of smokeless tobacco users. The utility of these assays to provide information about carcinogen dose, delineation of exposed vs. non-exposed individuals, and carcinogen metabolism in humans is discussed. NNAL and NNAL-Gluc are exceptionally useful biomarkers because they are derived from a carcinogen- 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK)- that is specific to tobacco products. The NNAL assay has high sensitivity and specificity, which are particularly important for studies on ETS exposure. Other useful assays that have been widely applied involve quantitation of 1-HOP and tt-MA. Urinary carcinogen metabolite biomarkers will be critical components of future studies on tobacco and human cancer, particularly with respect to new tobacco products and strategies for harm reduction, the role of metabolic polymorphisms in cancer, and further evaluation of human carcinogen exposure from ETS.

Analysis of N- and O-glucuronides of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) in human urine.

4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) is a tobacco-specific lung carcinogen which may play an important role as a cause of lung cancer in smokers. NNK is extensively metabolized to 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), which like NNK is a potent pulmonary carcinogen. NNAL in turn is glucuronidated, and both NNAL and its glucuronides are excreted in human urine. Previous studies have clearly demonstrated the presence in human urine of 4-(methylnitrosamino)-1-(3-pyridyl)-1-(O-beta-D-glucopyranuronosyl)butane (NNAL-O-Gluc), but did not exclude the presence of 4-(methylnitrosamino)-1-(3-pyridyl-N-beta-D-glucopyranuronosyl)-1-butanolonium inner salt (NNAL-N-Gluc). In this study, we quantified NNAL, NNAL-N-Gluc, and NNAL-O-Gluc in the urine of smokers, snuff-dippers, and people who used the oral tobacco product "toombak". The presence of NNAL-N-Gluc in the urine of toombak users was confirmed by LC-ESI-MS/MS. In smokers' urine, NNAL-N-Gluc, NNAL-O-Gluc, and NNAL comprised (mean +/- SD) 26.5 +/- 6.2, 32.1 +/- 17.6, and 41.4 +/- 16.6%, respectively, of total NNAL. In snuff-dippers' urine, the corresponding figures were 13.6 +/- 5.1, 46.6 +/- 11.7, and 36.6 +/- 9.3%. NNAL-N-Gluc comprised 50 +/- 25% of total glucuronidated NNAL in smokers and 24 +/- 12% in snuff-dippers. This difference was significant (P = 0.01), suggesting that smoking induces glucuronidation of NNAL. The results of this study demonstrate that NNAL-N-Gluc contributes substantially to NNAL-glucuronides in human urine. These results are important for a clearer understanding of mechanisms of detoxification of NNK in humans.

METABOLIC POLYMORPHISMS AND URINARY BIOMARKERS IN SUBJECTS WITH LOW BENZENE EXPOSURE

The effect of some common metabolic polymorphisms on the rate of trans,trans -muconic acid (TMA) and S -phenylmercapturic acid (SPMA) excretion was investigated in 169 policemen exposed to low benzene levels (<10 µg/m 3 ) during the work shift. End-shift urinary concentrations of TMA and SPMA, normalized to unmetabolized blood benzene concentration, were used as indicators of individual metabolic capacity. CYP2E1, NQO1, GSTM1, and GSTT1 polymorphisms were analyzed in all subjects by polymerase chain reaction (PCR)-restriction fragment length (RFL). The results obtained show significantly elevated levels of TMA and SPMA in urine of smokers compared to nonsmokers, whereas no correlation with environmental benzene was observed. TMA/blood benzene ratio was partially modulated by glutathione S -transferase (GST) genotypes, with significantly higher values in null individuals (GSTM1 and GSTT1 combined). However, a greater fraction of total variance of TMA/blood benzene in the study population was explained by other independent variables, that is, season of sampling, smoking habits, and gender. Variance in SPMA/blood benzene ratio was only associated with smoking and occupation, whereas no significant role was observed for the metabolic polymorphisms considered. These results suggest that in a population exposed to very low benzene concentrations, urinary TMA and SPMA levels are affected to a limited extent by metabolic polymorphisms, whereas other factors, such as gender, lifestyle, or other confounders, may account for a larger fraction of the interindividual variability of these biomarkers.

Biomonitoring of environmental tobacco smoke (ETS)-related exposure to 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK)

The exposure of non-smokers to the tobacco-specific N-nitrosamine 4-(N-methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), a rodent lung carcinogen, was determined in the air of various indoor environments as well as by biomonitoring of non-smokers exposed to environmental tobacco smoke (ETS) under real-life conditions using the urinary NNK metabolites 4-(N-methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) and [4-(N-methylnitrosamino)-1-(3-pyridyl)but-1-yl]-beta-O-D-glucosiduronic acid (NNAL-Gluc). NNK was not detectable (<0.5 ng m-3) in 11 rooms in which smoking did not occur. The mean NNK concentration in 19 rooms in which smoking took place was 17.5 (2.4-50.0) ng m-3. The NNK levels significantly correlated with the nicotine levels (r=0.856; p< 0.0001). Of the 29 non-smokers investigated, 12 exhibited no detectable NNAL and NNAL-Gluc excretion (<3 pmol day) in their urine. The mean urinary excretion of NNAL and NNAL-Gluc of the 17 remaining non-smokers was 20.3 (<3-63.2) and 22.9 (<3-90.0) pmol day-1, respectively. Total NNAL excretion (NNAL+NNAL-Gluc) in all non-smokers investigated significantly correlated with the amount of nicotine on personal samplers worn during the week prior to urine collection (r=0.88; <0.0001) and with the urinary cotinine levels (r=0.40; p=0.038). No correlation was found between NNAL excretion and the reported extent of ETS exposure. Average total NNAL excretion in the non-smokers with detectable NNAL levels was 74 times less than in 20 smokers who were also investigated. The cotinine/total NNAL ratios in urine of smokers (9900) and non-smokers (9300) were similar. This appears to be at variance with the ratios of the corresponding precursors (nicotine/NNK) in mainstream smoke (16400) and ETS (1000). Possible reasons for this discrepancy are discussed. The possible role of NNK as a lung carcinogen in non-smokers is unclear, especially since NNK exposure in non-smokers is several orders of magnitude lower than the ordinary exposure to exogenous and endogenous N-nitrosamines and the role of NNK as a human lung carcinogen is not fully understood.

Oxidants and antioxidants as therapeutic targets in chronic obstructive pulmonary disease.

There is now increasing evidence that an oxidant/antioxidant imbalance, in favor of oxidants, occurs in chronic obstructive pulmonary disease (COPD). Evidence is also accumulating that oxidative stress is a critical event in the pathogenesis of this condition. A large number of studies have demonstrated an increased oxidant burden and consequently increased markers of oxidative stress in the airspaces, breath, blood, and urine of smokers and of patients with COPD. There are several events related to oxidative stress, which are important in the pathogenesis of COPD. These include oxidative inactivation of antiproteinases, airspace epithelial injury, increased sequestration of neutrophils in the pulmonary microvasculature, and gene expression of proinflammatory mediators. Oxidative processes have a fundamental role in the inflammation of smokers and patients with COPD, through redox-sensitive transcription factors such as NF-kappaB and AP-1, which regulate the genes for proinflammatory mediators and protective mechanisms, such as antioxidant gene expression. In addition to the oxidative stress produced by cigarette smoking, dietary deficiency in antioxidants is related to the development of airflow limitation, and hence dietary supplementation may be a beneficial therapeutic intervention in this condition. The use of antioxidants with good bioavailability or molecules that have antioxidant enzyme activity may be treatments that not only protect against the direct injurious effects of oxidants, but may fundamentally alter the inflammatory events that are thought to play an important part in the pathogenesis of COPD. MacNee W, Rahman I. Oxidants and antioxidants as therapeutic targets in chronic obstructive pulmonary disease.

Extraction and high-performance liquid chromatographic separation of selected pyrene and benzo[ a]pyrene sulfates and glucuronides: preliminary application to the analysis of smokers’ urine

In the study of the complex mixture of urinary metabolites derived from polycyclic aromatic hydrocarbon compounds, it is desirable to simplify the analysis through separation of classes of compounds. We have developed a liquid chromatography (LC) method for the separation of selected sulfate and glucuronide conjugate isomers derived from hydroxybenzo[ a]pyrenes (OH-BaP) and hydroxypyrenes. This LC method was utilized in the preliminary analysis of the urine of smokers by combining it with an extraction technique employing tetra-n-butyl-ammonium ion as a coupling agent to generate a 1:1 complex, extractable in chloroform at low pH prior to LC analysis.

Profile of urinary phenanthrene metabolites in smokers and non-smokers

Phenanthrene metabolites (phenols and dihydrodiols) and 1-hydroxypyrene excreted in the 24-h urine of smokers, non-smokers and lung cancer patients, who after heavy smoking became light smokers, were determined and compared. In contrast to 1- hydroxypyrene, no significant differences of the absolute amounts of phenanthrene metabolites were found between smokers and non-smokers. A ratio phenanthrene metabolites/l-hydroxypyrene of 10.4 was observed for non-smokers and 9.9 for lung cancer patients, but 4.2 for smokers. Significantly different ratios for the regiospecific oxidation of phenanthrene were found for smokers when compared with non-smokers (1,2-oxidation vs 3,4-oxidation was 1.45 in the case of smokers, but 2.34 in the case of non-smokers) indicating a cigarette smoke - but not PAH - caused induction of CYP 1A2 in smokers. As a consequence of the degree of PAH exposure the ratio dihydrodiols/phenols depends on the total amount of metabolites excreted. Phenols predominate, equally in smokers and non-smokers after low exposure, while dihydrodiols become more prominent in highly exposed persons (coke plant workers). Both (i) the regiospecific oxidation of PAH and (ii) the ratio of dihydrodiol vs phenol formation may be recognized from the urinary phenanthrene metabolite profile. This pattern mirrors the enzymatic status (balance of the CYP isoforms and epoxide hydrolase) in individuals. Accordingly, more detailed information may be obtained from the urinary metabolite pattern than from 1- hydroxypyrene, commonly used in PAH biomonitoring.

Fast screening method for the profile analysis of polycyclic aromatic hydrocarbon metabolites in urine using derivatisation–solid-phase microextraction

A method for the qualitative analysis of various metabolites of naphthalene, phenanthrene and pyrene is presented. The method uses SPME with a 85-μm polyacrylate fibre for extraction, headspace silylation with BSTFA without any catalyst for on-fibre derivatisation and GC–MS in the SIR mode for separation and detection. The suitability of the method for profile analysis of PAH metabolites is shown by analysing a smokers urine after enzymatic cleavage (and additionally after spiking with the target analytes) and spiked water. The method exhibits satisfactory separation of all investigated metabolites and no interferences due to matrix peaks.

Increasing urinary cotinine concentrations at elevated temperatures: the role of conjugated metabolites

The presence of cotinine, a nicotine metabolite, in urine above a specified cutoff concentration is commonly used to distinguish smokers from nonsmokers, as in smoking cessation studies. A stability study of cotinine in urine was carried out after questions arose concerning analyte stability at elevated storage and shipment temperatures. Aliquots from a smokers urine pool were stored at 5, 25, 40, 50 and 60°C for 30 days. Another set of aliquots, obtained by diluting the smokers pool 1:1 with nonsmokers urine, were stored under the same conditions. Free cotinine levels, determined by a stability-indicating gas chromatographic/mass spectrometric (GC/MS) assay, increased over the 30-day period at higher storage temperatures. Cotinine concentrations in the aliquots stored at 60°C, for example, nearly doubled over 30 days (1301 to 2476 ng/ml), with similar proportional increases observed in the aliquots diluted with nonsmokers urine. Since cotinine can be excreted to a large degree as cotinine- N-glucuronide, this conjugated metabolite was determined by an indirect method. As the storage temperature increased, the free/conjugated cotinine ratio dramatically increased, pointing to cotinine- N-glucuronide as the source of the additional free cotinine at the higher storage temperatures. The results of this study are of much practical importance, since urine samples with free cotinine concentrations just below a cutoff value may become positive for smoking status if suitably low temperatures cannot be maintained during sample handling and shipment.

N-nitroso compounds and man

Based on recent analytical data, total human exogenous exposure to N-nitrosamines is estimated to be 1.10 mumol/day; the major exposure sources are the diet (0.79 mumol/may, 80-120 micrograms/day; 72%), occupational exposure (0.15-0.30 mumol/day; 25%), cigarette smoking (0.02 mumol/day, 3.4 micrograms/day; 2%), and miscellaneous minor sources, including pharmaceutical products, cosmetics, indoor and outdoor air (0.001 mumol/day, 0.1 micrograms/day; 1%). Excretion of apparent total N-nitroso compounds (ATNC) in healthy adults is estimated to be 1.30 +/- 1.05 mumol/day in urine and between 1.56 +/- 1.56 and 3.17 +/- 2.58 mumol/day in faeces. The excretion of volatile N-nitrosamines (N-nitrosodimethylamine), and N-nitrosamine acids and their derivatives (N-nitrososarcosine, N-nitrosoproline, N-nitrosothiazolidine-4-carboxylic acid and N-nitroso-2-methylthiazoline-4-carboxylic acid) accounts for approximately 0.03% and 16.0% of urinary ATNC, respectively. 4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanol and its O-glucuronide conjugate, two metabolites of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone present in urine of smokers, account for 0.2% of the urinary ATNC response; < 1.5 of the excretion of currently identified N-nitroso compounds in urine. The remaining N-nitroso compounds excreted in urine and those present in faeces are still unidentified. A crude mass balance between exogenous exposure and excretion in urine and faeces indicates that 45-75% of the total human exposure to N-nitroso compounds results through in vivo formation.

Use of Salmonella typhimurium TA 98, YG 1024 and YG 1021 and deconjugating enzymes for evaluating the mutagenicity from smokers' urine

Four smokers were chosen for their different smoking habits, and their declared cigarette consumption confirmed by urinary measurement of nicotine and its metabolites. The promutagenicity of their urine was evaluated by the Ames test, modified according to Kado et al. (Mutation Res., 31 (1983) 25–32) after extraction on XAD2 Amberlite resin. The different Salmonella typhimurium strains TA 98, YG 1021 and YG 1024 were compared to determine the presence of amino aromatic compounds in the urine of smokers of blond and black tobacco. The strain YG 1024 shows higher mutagenicity than TA 98 for extracts from the smoker's urine and more particularly from black tobacco smokers. In addition, the pretreatment of urine by external enzymatic systems (β-glucuronidase or arylsulfatase) reveals the presence in the urine of glucurono- and sulfoconjugated forms of promutagens, including amino aromatic compounds.

Interference of Nicotine Metabolites in Cotinine Determination by RIA

Cotinine (COT), a major metabolite of nicotine (NIC), has been used as a biomarker in many studies of active and passive smoking (1)(2)(3)(4). Methods of analysis for COT in biological fluids include gas chromatography, gas chromatography–mass spectrometry, HPLC, HPLC–mass spectrometry, and immunoassays (5)(6)(7)(8)(9)(10). In particular, the RIA developed for COT and NIC by Langone and Van Vunakis (11) was the method reported by the International Agency for Research on Cancer (IARC) for determination of these analytes in several biological matrices (12)(13). In the description of the assay (11), no mention was made about cross-reactivity of anti-NIC and anti-COT antibodies with trans -3′-hydroxycotinine (THOC) and cotinine glucuronide (COT-G). Indeed, these two compounds were found to be the most abundant metabolites of NIC in urine of smokers (14). Other studies, however, have investigated the cross-reactivity of the anti-COT antiserum with THOC and other metabolites in an ELISA (15). The objective of this study was to investigate the cross-reactivity of the anti-COT and anti-NIC antisera with THOC and COT-G in the RIA (11)(12)(13). We also used urine samples from active and passive smokers to compare data from the RIA with data obtained by HPLC. The rabbit antiserum, (−)[3H]COT, and (−)[3H]NIC were …

Heavy cigarette smokers show higher mutagenicity in urine

We examined the mutagenicity of cigarette smoker's urine in 32 healthy male cigarette smoker and 37 healthy male non-smoker. Twenty-four-hour urine specimens were subjected to blue rayon extraction which selectively adsorb polycyclic compounds, after which the elutions were fractionated by carboxymethyl cellulose column chromatography for removing antimutagenic compounds. The mutagens were measured by using an S9-mediated Salmonella mutagenicity test on strain TA98. Compared with those with non-smokers, smokers' urine showed a significantly higher urinary level of mutagens in the acid-elutable and in the sum of all chromatography fractions. A similar tendency was also seen in the alkali-elutable fraction. The subjects were classified into three groups according to the number of smoked cigarettes. Heavy smokers, who smoked more than 20 cigarettes per day, showed a significantly higher urinary level of mutagens than both non-smokers and light smokers especially in the acid-elutable and in the sum of all chromatography fractions. Our findings suggest that smokers are exposed to a great amount of polycyclic carcinogens and mutagens by cigarette smoking. These results also suggest that urinary level of mutagens measured by using blue rayon extraction combined with carboxymethyl cellulose chromatography could be a good index for estimating the exposure to carcinogens and mutagens such as polycyclic compounds.