Hair nicotine/cotinine concentrations as a method of monitoring exposure to tobacco smoke among infants and adults

In this pilot study, we examined the validity and usefulness of hair nicotine-cotinine evaluation as a biomarker of monitoring exposure to tobacco. Head hair samples were collected from 22 infants (<2 years of age) and 44 adults with different exposures to tobacco (through either active or passive smoking) and analyzed by liquid chromatography-mass spectrometry (LC-MS) for nicotine and cotinine. Hair samples were divided into three groups, infants, passive smoker adults and active smoker adults, and into eight subgroups according to the degree of exposure. The limit of quantification (LOQ) was 0.1 ng/mg for nicotine and 0.05 ng/mg for cotinine. Mean recovery was 69.15% for nicotine and 72.08% for cotinine. The within- and between-day precision for cotinine and nicotine was calculated at different concentrations. Moreover, hair nicotine and cotinine concentrations were highly correlated among adult active smokers (R (2) = 0.710, p < 0.001), among adult nonsmokers exposed to secondhand smoke (SHS; R (2) = 0.729, p < 0.001) and among infants (R (2) = 0.538, p = 0.01). Among the infants exposed to SHS from both parents the noted correlations were even stronger (R (2) = 0.835, p = 0.02). The above results identify the use of hair samples as an effective method for assessing exposure to tobacco, with a high association between nicotine and cotinine especially among infants heavily exposed to SHS.     

Assessment of prenatal smoke exposure by determining nicotine and its metabolites in maternal and neonatal urine

Urine specimens were collected from 75 pregnant women before childbirth and from their newborns within 48 postnatal hours. A high-performance liquid chromatography (HPLC) method was used to determine urinary nicotine and its metabolites, cotinine and trans-3'-hydroxycotinine (OH-cotinine) to objectivise prenatal smoke exposure. Using the sum of nicotine metabolites as a marker, 34 women were classed as not exposed to smoke ( < 15 nmol/l), 18 as passive smokers (15-400 nmol/l), and 23 as active smokers ( > 400 nmol/1). The newborns of active smokers exhibited significantly (P < 0.001) higher nicotine metabolite concentrations than did those of either non-exposed women or passive smokers. A close correlation was found to exist between maternal and neonatal nicotine and cotinine concentrations (r=0.8968 and r=0.9205, respectively). For OH-cotinine, this correlation was particularly close when maternal, but not neonatal, OH - cotinine was adjusted to creatinine (r=0.9792). The neonatal/maternal urine concentration ratios for cotinine and OH-cotinine were noted to not significantly depend on the time of postpartal urine collection. Within the first two postnatal days, the extent of current prenatal smoke exposure attributable to active smoking of the mother was best reflected by the urinary concentrations of cotinine plus OH-cotinine without adjustment to creatinine.     

Smoking and passive smoking during pregnancy and early infancy: effects on birth weight, lactation period, and cotinine concentrations in mother's milk and infant's urine

The extent of smoke exposure via mother's milk and passive smoking was investigated in a prospective, longitudinal matched-pair study by comparison between children, whose mothers smoked substantially throughout pregnancy and nursing period and children whose mothers did not smoke. Our preliminary results show that not only infants of smoking mothers but also those of smoking fathers show reduction of birth weight. Smoking mothers weaned their babies earlier than non-smokers. Cotinine concentrations in breast milk depended on the number of cigarettes smoked. The highest urinary excretion of cotinine (as expressed by ng cotinine/mg creatinine ratios) were observed in infants fully breast-fed by smoking mothers. After weaning the values were in the same range as those of formula-fed infants of smoking mothers (exposed to passive smoking only). In the group of non-smokers only small or undetectable amounts of cotinine were found. Thus it is demonstrated that both nursing and--to a lower degree--passive smoking contribute to the exposure of infants to nicotine and its metabolite cotinine.     

Reference values for hair cotinine as a biomarker of active and passive smoking in women of reproductive age, pregnant women, children, and neonates: systematic review and meta-analysis

Exposure to environmental tobacco smoke (ETS) is most often estimated using questionnaires, but they are unreliable. Biomarkers can provide valid information on ETS exposure, the preferred biomarker being cotinine. However, no reference range of hair cotinine exists to distinguish among active, passive, and unexposed nonsmokers. This study identifies cutoffs to validate cotinine as a marker for exposure to ETS. Data were obtained from six databases (four US, one Canada, one France). Active smoking and exposure to ETS were measured in the hair of women of reproductive age, pregnant women, their children, and neonates. Subjects were classified into active smokers, passively exposed to ETS, and unexposed nonsmokers. A total of 1746 cases were available for analysis. For active smokers, mean hair cotinine concentrations (95% confidence interval) were 2.3 to 3.1 ng/mg for nonpregnant women and 1.5 to 1.9 ng/mg for pregnant women. In the group of passive smokers, mean hair cotinine concentrations were 0.5 to 0.7 ng/mg for nonpregnant women, 0.04 to 0.09 ng/mg for pregnant women, 0.9 to 1.1 for children, and 1.2 to 1.7 for neonates. Among unexposed nonsmokers, mean hair cotinine was 0.2 to 0.4 ng/mg in nonpregnant women, 0.06 to 0.09 ng/mg in pregnant women, and 0.3 to 0.4 ng/mg in children. Cutoff values for hair cotinine were established to distinguish active smokers from passive or unexposed (0.8 ng/mg for nonpregnant women and 0.2 ng/mg for pregnant women). A cutoff value of 0.2 ng/mg was accurate in discriminating between exposed children and unexposed. These new values should facilitate clinical diagnosis of active and passive exposure to tobacco smoke. Such diagnosis is critical in pregnancy and in a large number of tobacco-induced medical conditions.     

Passive smoke exposure in infants and children with respiratory tract diseases.

1. The adverse effect of passive smoke exposure on the respiratory tract, particularly in infants and children, is not an issue of dispute. It was the objective of this study to analyse the extent and the intensity of passive smoke exposure in infants and children with respiratory tract diseases, and compare the information obtained with parents' subjective assessment. At the time of admission to the hospital, the parents of 295 infants and children (aged 1 month to 11 years) were questioned by the physician as to the smoking habits in the families' homes. An HPLC method was employed to determine simultaneously nicotine, cotinine and trans-3'-hydroxycotinine in the children's urine. 2. The sum of the nicotine metabolites turned out to be a sensitive marker in determining passive smoke exposure. Measurements revealed passive smoke exposure in 66% of the children, the frequency in younger children being significantly (P < 0.001) higher than in children over 5 years (84% vs 52%). The average concentration of nicotine metabolites in younger passive smokers was significantly (P < 0.001) higher when compared to the older ones (193 nmol/l vs 86 nmol/l). Forty-nine per cent of the parents assessed that their children had experienced passive smoke exposure, and another 10% emphasised that they only smoked in the absence of their child. In children with cystic fibrosis and bronchial asthma, the number of passive smokers as assessed by their parents were lower by 65% and 29% respectively when compared to the findings obtained from measurements. In children without respiratory diseases, the difference was as little as 18%. 3. Parents when questioned in conjunction with an illness of their children, tended to understate, or even withhold the truth about, passive smoke exposure. Therefore, reliable information on passive smoke exposure of patients can only be obtained through objective measurements.     

Estimation of smoke dosage and mortality of non-smokers from environmental tobacco smoke

This paper shows how biochemical markers can be used to estimate smoke intake from passive smoking to complement epidemiological studies on the health risks and mortality to non-smokers. Using data from slow nicotine infusions given over 1 h, we estimated that the nicotine intake from passive smoking averages about 0.014 mg/h among urban non-smokers leading their usual daily lives. This compares with 0.23 mg/h in a smoke-filled public house, 0.36 mg/h during maximum exposure in an unventilated room, and 1.0 to 1.4 mg nicotine per cigarette taken in by active smokers. Data from several studies on urinary nicotine concentrations and those of cotinine in blood, urine and saliva were collated. The results show that the concentrations in non-smokers averaged about 0.7% (for nicotine) and 0.6% (for cotinine) of the levels found in smokers. From this we estimate that the mortality from passive smoking is about 1000 non-smokers per year in Britain and about 4000 per year in the United States, assuming that the relation of dose to risk is linear.     

Hair nicotine:cotinine metabolic ratio in pregnant women: a new method to study metabolism in late pregnancy

A large number of smoking women cannot quit their habit when they become pregnant. Preliminary evidence suggests an enhanced nicotine clearance rate in late pregnancy. To evaluate the change in nicotine metabolism in late pregnancy, we undertook a prospective cohort study of a large, diverse group of pregnant women recruited from four Montreal maternal hospitals. Smoking histories were obtained by structured questionnaires administered at 24 to 26 weeks of gestation and postpartum. Hair concentrations of nicotine and cotinine were measured by immunoassays for each trimester based on sectioning the hair and assuming average hair growth of 1 cm per month. A strong correlation was observed between average number of cigarettes smoked per day and hair nicotine and cotinine in all three trimesters. A significant decrease in hair nicotine was observed among steady smokers from the first to third trimester paralleled by a significant increase in hair cotinine. The ratio of hair nicotine:cotinine decreased significantly from the first to the third trimester. Increased nicotine metabolism in late pregnancy results in lower systemic exposure to nicotine. This phenomenon may explain why many pregnant women feel the urge to continue smoking and why standard-dose nicotine replacement therapy has not been effective in reducing smoking during pregnancy in several clinical trials.     

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.     

Implications of nicotine detected in autopsy cases of newborn babies and infants from the perspective of social medicine

Smoking by pregnant and parturient women is generally suspected to increase nicotine levels in fetal and infant blood. Supportive data of nicotine levels in infants is, however, inadequate. We investigated blood and muscle nicotine and cotinine levels in 14 autopsy cases of newborn babies and infants using gas chromatography. Among the 14 cases investigated, nicotine or cotinine was detected in six cases (42.9%). In each of these six cases, the mother was a smoker. Route of exposure to nicotine originating from smoking was transplacental in three cases, via breast milk in one case and secondhand smoke in two cases. Nicotine and cotinine levels in blood from the two cases with placental exposure were 10.6-84.4 ng/ml and 20.3-183 ng/ml, and levels in muscle from one case were 43.9 ng/g and 308 ng/g, respectively. Nicotine and cotinine levels in blood from exposure via breast milk were 19.1 ng/ml and 87.1 ng/ml, and from secondhand smoke were 0 ng/ml and 14.6-20.1 ng/ml. Mean concentrations of blood nicotine and cotinine in 68 autopsy cases of adult habitual smokers were 30.0 ng/ml and 247 ng/ml. Our data for nicotine and cotinine levels in infant blood seem to indicate that some infants who are born and develop under exposure to smoking by family members, particularly the mother, may show high nicotine levels in blood and experience possible health risks.     

Nicotine exposure can be detected in cerebrospinal fluid of active and passive smokers

A simple, rapid and sensitive liquid chromatography/mass spectrometry (LC/MS) method has been utilized for the quantitative determination of nicotine and its major metabolite cotinine (COT) in human cerebrospinal fluid (CSF) of active and passive smokers. CSF samples from 18 smokers, 15 non-smokers, 15 children, 15 infants, and 9 neonatal were analyzed for nicotine (NIC) and cotinine content. Cotinine levels in the CSF of smokers ranged from 27.3 to 457.1 ng/ml, whereas nicotine levels were considerably lower (6.0–215.1 ng/ml). Cotinine could be detected in 4 of the 15 CSF samples from non-smokers (3.5–30.4 ng/ml), and a few other passive smokers, including neonates from smoking mothers (15.6–81.1 ng/ml). The concentrations of cotinine in CSF samples suggests that nicotine easily passes into the CSF, which makes it an excellent CSF marker for tobacco-smoke exposure.     

Total cotinine in plasma: a stable biomarker for exposure to tobacco smoke

Determination of plasma cotinine concentration is the predominant assay employed to quantify smoking and exposure to environmental tobacco smoke in epidemiological studies. However, cotinine is biotransformed into secondary metabolites. This pilot study determined plasma concentrations of cotinine, cotinine glucuronide, 3-hydroxycotinine, and 3-hydroxycotinine glucuronide. Total cotinine concentration was determined by summation of all four metabolites. The goals of this study were (1) to explore the stability and validity of total cotinine concentration as a measure of tobacco smoking and as a measure of exposure to environmental tobacco smoke in nonsmokers, (2) to explore the stability of plasma concentrations of each of the four nicotine metabolites in smokers by performing a.m. and p.m. measures, and (3) to explore the stability of indices of glucuronidation as measures of possible markers for enzymatic activity. The subject sample included 76 white volunteers (32% smokers and 68% nonsmokers). Plasma total cotinine concentration appeared to be very stable, suggesting that total cotinine concentration may be a good measure for epidemiological studies employing a single plasma sample. Moreover, plasma total cotinine concentration also reflected exposure to environmental tobacco smoke more accurately than did plasma cotinine concentration, which would have not identified 27% of passive smokers. 3-Hydroxycotinine glucuronide and 3-hydroxycotinine plasma concentrations were almost as stable as cotinine concentrations. However, cotinine glucuronide and its indices of glucuronidation were unstable, suggesting that cotinine glucuronide undergoes deconjugation. New studies of total cotinine in plasma using more than two blood collections during the day are needed to definitively establish that it is a stable biomarker for epidemiological studies.     

Urinary hydroxyproline excretion in smokers, non-smokers and passive smokers

The hydroxyproline/creatinine ratio in urine was investigated in 200 cigarette smokers, 199 pipe and/or cigar smokers and 24 non-smokers. For cigarette smokers a statistically significant positive correlation is found between this ratio and daily cigarette consumption, COHb, serum cotinine and nicotine excretion in urine. This smoking-related increase in the hydroxyproline/creatinine ratio is, for the most part or completely, due to the fact that creatinine urine concentrations inversely correlate with the smoke uptake variables. Neither pipe and/or cigar smoking nor passive smoke exposure of non-smokers seem to affect the hydroxyproline/creatinine ratio. A seasonal influence is found in these studies as well as in two experiments with limited numbers of subjects: the hydroxyproline/creatinine ratio is higher in winter than in summer for both smokers and non-smokers. Our data do not favour the idea that measuring hydroxyproline/creatinine ratios in urine is an accurate method of investigating early effects of smoking, passive smoking and air pollution in man.     

A rapid gas-liquid chromatographic method for the determination of cotinine and nicotine in biological fluids

A rapid method is described for the simultaneous measurement of nicotine and cotinine in biological fluids using capillary column gas-liquid chromatography. Using 100 microliters sample volume the lower limit of detection for both nicotine and cotinine was 100 pg mL-1, allowing the method to be used for the measurement of these compounds in both smokers and non-smokers. The extraction time is 3 min per sample, and by using multi-pipetting and vortexing systems 250 samples can be extracted per day. The average coefficient of variation over the nicotine range 1.0 to 100 ng mL-1 was 3.9% and for cotinine over the range 1.0 to 1000 ng mL-1 was 2.2%. Saliva cotinine concentrations were quantitatively related to passive exposure to parental smoking in a population study of 1118 non-smoking schoolchildren.     

Comparison of serum and salivary cotinine measurements by a sensitive high-performance liquid chromatography-tandem mass spectrometry method as an indicator of exposure to tobacco smoke among smokers and nonsmokers

Exposure to tobacco smoke, both from active smoking and from passive exposure to environmental tobacco smoke, can be monitored by measuring cotinine, a metabolite of nicotine, in a variety of biological sources including blood, urine, and saliva. Previously, a sensitive atmospheric-pressure ionization, tandem mass spectrometric (LC-API-MS-MS) method for cotinine measurements in serum was developed in support of a large, recurrent national epidemiologic investigation. The current study examined the application of this LC-API-MS-MS method to both serum and saliva cotinine measurements in a group of 200 healthy adults, including both smokers and nonsmokers. The primary objective of this study was to evaluate the relationship between serum and saliva cotinine concentrations to facilitate the linking of results from epidemiologic studies using salivary cotinine measurements to existing national data based on serum cotinine analyses. The results indicate that a simple, linear relationship can be developed to describe serum and saliva cotinine concentrations in an individual, and the expression describing this relationship can be used to estimate with reasonable accuracy (approximately +/- 10%) the serum cotinine concentration in an individual given his or her salivary cotinine result. It was further confirmed that saliva cotinine samples are generally quite stable during storage after collection, even at ambient temperatures, and this sample matrix appears to be well-suited to the requirements of many epidemiologic investigations.     

Environmental tobacco smoke at home and in public places prior to smoking ban enforcement: Assessment by hair analysis in a population of young adult students

Despite inititatives to reduce tobacco consumption, smoking remains a primary cause of death for both smokers and nonsmokers exposed to environmental tobacco smoke (ETS). The characteristics of some specific groups can make them more exposed to ETS or limit the benefit of prevention measures. This study investigated determinants of ETS in a population of young adult students, considered at higher risk of exposure due to their specific lifestyle. This cross-sectional study involved 90 students aged 20 ± 1.7 years, from the University of Luxembourg, prior to the smoking ban enforcement in public places in the country. Participants reported their tobacco consumption and exposure to ETS at home and/or in public places, and provided a hair sample analyzed for nicotine and cotinine. Nicotine and cotinine were significantly higher in smokers than in nonsmokers' hair in general (median: 2.6 vs. 0.9 ng/mg and 87.1 vs. 22.5 pg/mg respectively). However, nonsmokers exposed to ETS at home and in public places had comparable concentrations to smokers (nic = 2.2 ng/mg; cot = 56.2 pg/mg), whereas unexposed nonsmokers presented significantly lower values (nic = 0.4 ng/mg, cot = 8.5 pg/mg). Nonsmokers exposed to ETS only at home presented higher values than nonsmokers only exposed in public places (nic: 1.3 vs. 0.8 ng/mg, cot: 70.4 vs. 15.0 pg/mg). The study shows the widespread exposure to ETS in this population, the importance of exposure assessment, and the relevance of hair analysis for this purpose. Results suggest that ETS can lead to equivalent exposure to active smoking and that exposure at home can highly contribute to ETS, which is not solved by smoking ban in public places.     

Household smoking behavior and ETS exposure among children with asthma in low-income,minority households

Environmental tobacco smoke (ETS) exposure was measured among 242 children with asthma who live in homes where at least one person smokes. Subjects were identified through clinics, schools, community agencies, and hospitals serving low-income, medically underserved communities in Los Angeles. Parents were surveyed about smoking behaviors in the household, children's ETS exposure, and attitudes towards smoking and smoking behavior change. Validation measures included urine cotinine for the child with asthma and passive air nicotine monitors placed in the subjects' homes. Overall reported levels of household smoking and ETS exposure were low, with a significant amount of household smoking taking place outside rather than inside the home. Over 47% of the respondents reported absolute restrictions against smoking in the home, and these restrictions were associated with lower reported levels of smoking, ETS exposure, and air nicotine and urine cotinine concentrations.     

Assessing the accuracy of self‐reported smoking status and impact of passive smoke exposure among pregnant Aboriginal and Torres Strait Islander women using cotinine biochemical validation

Introduction and Aims. A significant level of misreport or error occurs during questionnaire‐based assessment of smoking behaviour. Failure to measure environmental tobacco smoke, and participant's inclination to under‐report their smoking raise questions as to the accuracy of assessment. In order to establish an estimation of the possible error associated with such assessment, the accuracy of self‐reported smoking status among a group of pregnant Aboriginal and Torres Strait Islander women was examined. Design and Methods. Women attending two Aboriginal Medical services in Far North Queensland for antenatal care were invited to participate. Women completed an interviewer assisted questionnaire relating to their smoking status and a 24 h diary of their exposure to nicotine and consumption of alcohol. Urine samples were analysed for cotinine using an Enzyme Linked Immunosorbent Assay. Results. Cotinine analysis indicated that 17% of women who reported that they were non‐smokers were likely to have misreported this status, or be exposed to high levels of passive smoke. The only significant predictors of cotinine level were self‐reported nicotine exposure (including passive smoke) and number of cigarettes smoked in the previous 24 h. Other individual and environmental variables had no significant influence on cotinine level using this analysis technique. Discussion and Conclusions. The level of potential error in smoking assessment among this group was substantial. Exposure to environmental tobacco smoke might explain part of this error, but the reasons for misreport can only be speculated. This rate of misclassification should be taken into consideration in routine screening of antenatal women in primary health care.[Gilligan C, Sanson‐Fisher R, Eades S, Wenitong M, Panaretto K, D'Este C. Assessing the accuracy of self‐reported smoking status and impact of passive smoke exposure among pregnant Aboriginal and Torres Strait Islander women using cotinine biochemical validation. Drug Alcohol Rev 2009]     

TDM grand rounds: neonatal nicotine withdrawal syndrome in an infant prenatally and postnatally exposed to heavy cigarette smoke

A heavy smoking, lactating mother delivered a baby that exhibited spontaneous tremors, fluctuations of muscular rigidity, and opisthotonus at 48 hours of life. Although the symptoms did not disappear within the following days, they could be controlled by swaddling or wrapping the baby in a blanket. The absence of any other etiology generated a suspicion of prenatal exposure to heavy tobacco smoke and potential neonatal nicotine withdrawal syndrome. This diagnosis was supported by extremely high concentration of hair nicotine and cotinine in the infant's hair and in different segments of maternal hair. The presence of non-negligible amounts of nicotine and cotinine in breast milk confirmed that the mother did not quit smoking after delivery, despite her reports. The breast-fed newborn continued to have 3 to 4 crises of spontaneous tremors and alternant muscular rigidity per day for a month. More studies are needed to establish neonatal nicotine withdrawal.     

Time course of nicotine and cotinine incorporation into samples of nonsmokers' beard hair following a single dose of nicotine polacrilex

Hair nicotine and cotinine have been proposed as longer-term markers of exposure to secondhand smoke. In this study, we evaluated the rate and extent of nicotine and cotinine deposition into beard hair among six male nonsmokers following a single exposure to 4 mg of nicotine in Nicorette(?) (nicotine polacrilex) gum. We collected beard hair samples daily for 12 days following exposure and urine samples for 6 days after exposure. Using liquid chromatographic-tandem mass spectrometric analysis, we found that both nicotine and cotinine could be detected in beard samples within 24 h of the exposure and reached a maximum of about 71 pg nicotine and 47 pg cotinine/mg hair, respectively, within 1-2 days, followed by a gradual decline. Compared to beard hair concentrations, nicotine, cotinine, and hydroxycotinine were excreted in urine at much higher levels and also peaked on the day after exposure (mean ± SD urine cotinine = 300 ± 183 ng/mL). Our results confirmed that both nicotine and cotinine can be measured in beard hair samples following a single dose of nicotine. However, both the time-course and extent of deposition of these analytes in beard hair in this study differed from the results reported previously from a similar evaluation.     

Low level saliva cotinine determination and its application as a biomarker for environmental tobacco smoke exposure.

1. The determination of personal exposures to environmental tobacco smoke (ETS) and respirable suspended particles (RSP) for housewives, and fixed site monitoring of their homes, have been undertaken by these authors throughout Europe, South East Asia and Australia. Median 24 h time weighted average (TWA) concentrations for ETS particles and nicotine were found to be significantly higher for housewives living in smoking households compared with those living in nonsmoking households. For Europe, median TWA concentrations of 4.1 and <0.26 microg/ml for ETS particles and 0.63 and < 0.08 microg/m3 for nicotine were found for housewives living in smoking and nonsmoking households respectively. 2. In addition to the measurement of RSP, ETS particles and nicotine, saliva cotinine concentrations were determined using a radioimmunoassay method with a limit of quantitation of 1 ng/ml. Median saliva cotinine concentrations of 1.4 and <1 ng/ml were determined for European housewives living in smoking and nonsmoking households respectively, which reflected the poor limit of quantitation of this methodology. A chromatographic method utilising tandem mass-spectrometric detection was developed and validated for the determination of both cotinine and 3-hydroxycotinine, two of the main metabolites of nicotine, with lower limits of quantitation of 0.05 and 0.10 ng/ml respectively. This method was applied to samples collected from subjects with a known ETS exposure history and median cotinine concentrations of <0.05 ng/ml for self-reported unexposed nonsmokers, 0.65 ng/ml for nonsmokers reporting some ETS exposure and 1.28 ng/ml for nonsmokers living with smokers were found. 3. In conclusion, the measurement of RSP and ETS concentrations derived from personal or fixed site monitors for housewives may provide some indication of potential exposures to dependent children. The recent development and application of a highly sensitive assay for the determination of cotinine in saliva has provided evidence to suggest that concentrations determined at sub-nanogram levels may be used as a biomarker for ETS exposure. This improved methodology, coupled with non-invasive sampling for saliva, may be of significance when considering the application of cotinine as a biomarker for ETS exposure in children.