In vivo quantification and pharmacokinetic studies of cotinine in mice after smoke exposure by LC-MS/MS.

A sensitive analytical method was developed and validated for the quantification of cotinine in mouse plasma after exposure to smoke of 0.5, 1.0, and 1.5 commercially available cigarettes, using liquid chromatography tandem mass spectrometry. The method was validated over a linear concentration range of 0.075-20.0 ng/mL with the R2 value being higher than 0.99. Both the precision (coefficient of variation; %) and accuracy (relative error; %) were within acceptable criteria of <15%. The lower limit of quantification (LLOQ) for cotinine was 0.075 ng/mL with sufficient specificity, accuracy, and precision. Following exposure to 0.5, 1.0, and 1.5 cigarette smoke, it was observed that the AUC and the Cmax increased linearly as the doses increased. The pharmacokinetics of cotinine was found linear for the range of 0.5-1.5 commercial cigarette smoke. The quantification of the concentration of cotinine in mouse plasma after smoke exposure will facilitate future behavioral and toxicological experiments in animals and may prove useful in predicting cotinine levels in humans during smoking.

Simultaneous determination of plasma nicotine and cotinine by UHPLC-MS/MS in C57BL/6 mice and its application in a pharmacokinetic study.

Plasma concentrations of nicotine and its active metabolite cotinine are highly correlated with its biological effects. A UHPLC-MS/MS method was developed, validated and applied for nicotine and cotinine analysis in mice plasma. Chromatographic separation was achieved on a BEH HILIC column using acetonitrile (0.1% formic acid) and 10 mm ammonium formate as mobile phase. The gradient elution was performed at 0.4 mL/min with a run time of 3.6 min. The quantitative ion transition was m/z 163.1 > 130.0 for nicotine, m/z 177.1 > 80.0 for cotinine and m/z 167.1 > 134.0 for nicotine-D4 (internal standard, IS). For both nicotine and cotinine, the calibration range was 5-500 ng/mL with 5 ng/mL as the lower limit of quantitation, and the intra- and inter-day bias and imprecision were -4.61-12.00% and <11.12%. The IS normalized recovery was 90.62-98.95% for nicotine and 89.18-101.53% for cotinine, and the IS normalized matrix factor was 106.00-116.44% for nicotine and 100.34-109.85% for cotinine. Both nicotine and cotinine were stable under conventional storage conditions. The validated method has been applied to a pharmacokinetic study in mice to calculate the pharmacokinetic parameters for both analytes.

Effect of UGT2B10, UGT2B17, FMO3, and OCT2 genetic variation on nicotine and cotinine pharmacokinetics and smoking in African Americans.

OBJECTIVES: Nicotine metabolism rates differ considerably among individuals, even after controlling for variation in the major nicotine-metabolizing enzyme, CYP2A6. In this study, the impact of genetic variation in alternative metabolic enzymes and transporters on nicotine and cotinine (COT) pharmacokinetics and smoking was investigated. METHODS: We examined the impact of UGT2B10, UGT2B17, FMO3, NAT1, and OCT2 variation on pharmacokinetics and smoking (total nicotine equivalents and topography) before and after stratifying by CYP2A6 genotype in 60 African American (AA) smokers who received a simultaneous intravenous infusion of deuterium-labeled nicotine and COT. RESULTS: Variants in UGT2B10 and UGT2B17 were associated with urinary glucuronidation ratios (glucuronide/free substrate). UGT2B10 rs116294140 was associated with significant alterations in COT and modest alterations in nicotine pharmacokinetics. These alterations, however, were not sufficient to change nicotine intake or topography. Neither UGT2B10 rs61750900, UGT2B17*2, FMO3 rs2266782, nor NAT1 rs13253389 altered nicotine or COT pharmacokinetics among all individuals (n=60) or among individuals with reduced CYP2A6 activity (n=23). The organic cation transporter OCT2 rs316019 significantly increased nicotine and COT Cmax (P=0.005, 0.02, respectively) and decreased nicotine clearance (P=0.05). UGT2B10 rs116294140 had no significant impact on the plasma or urinary trans-3'-hydroxycotinine/COT ratio, commonly used as a biomarker of CYP2A6 activity. CONCLUSION: We found that polymorphisms in genes other than CYP2A6 represent minor sources of variation in nicotine pharmacokinetics, insufficient to alter smoking in AAs. The change in COT pharmacokinetics with UGT2B10 rs116294140 highlights the UGT2B10 gene as a source of variability in COT as a biomarker of tobacco exposure among AA smokers.

A novel delivery platform for therapeutic peptides.

Although many peptides have therapeutic effects against diverse disease, their short half-lives in vivo hurdle their application as drug candidates. To extend the short elimination half-lives of therapeutic peptides, we developed a novel delivery platform for therapeutic peptides using an anti-hapten antibody and its corresponding hapten. We selected cotinine because it is non-toxic, has a well-studied metabolism, and is physiologically absent. We conjugated WKYMVm-NH2, an anti-sepsis therapeutic peptide, to cotinine and showed that the conjugated peptide in complex with an anti-cotinine antibody has a significantly improved in vivo half-life while retaining its therapeutic efficacy. We suggest that this novel delivery platform for therapeutic peptides will be very useful to develop effective peptide therapeutics.

Pharmacokinetic and pharmacodynamics studies of nicotine after oral administration in mice: effects of methoxsalen, a CYP2A5/6 inhibitor.

INTRODUCTION: The use of novel oral nicotine delivery devices and compositions for human consumption and for animal research studies has been increasing in the last several years. METHODS: Studies were undertaken to examine whether the systemic administration of methoxsalen, an inhibitor of human CYP2A6 and mouse CYP2A5, would modulate nicotine pharmacokinetics and pharmacological effects (antinociception in the tail-flick, and hot-plate tests, and hypothermia) in male ICR mouse after acute oral nicotine administration. RESULTS: Administration of intra peritoneal (ip) methoxsalen significantly increased nicotine's Cmax, prolonged the plasma half-life (fourfold decrease) of nicotine, and increased its area under the curve (AUC) compared with ip vehicle treatment. Methoxsalen pretreatment prolonged the duration of nicotine-induced antinociception and hypothermia (15mg/kg, po) for periods up to 6- and 24-hr postnicotine administration, respectively. Additionally, methoxsalen potentiated nicotine-induced antinociception and hypothermia as evidenced by leftward shifts in nicotine's dose-response curve. Furthermore, this prolongation of nicotine's effects after methoxsalen was associated with a parallel prolongation of nicotine plasma levels in mice. These data strongly suggest that variation in the rates of nicotine metabolic inactivation substantially alter pharmacological effects of nicotine given orally. CONCLUSION: We have shown that the pharmacological effects of inhibiting nicotine's metabolism after oral administration in mice are profound. Our results suggest that inhibiting nicotine metabolism can be used to dramatically enhance nicotine's bioavailability and its resulting pharmacology, which further supports this inhibitory approach for clinical development of an oral nicotine replacement therapy.

Race, gender, and nicotine metabolism in adolescent smokers.

INTRODUCTION: Differences in the rate of nicotine metabolism between genders and different races have been hypothesized to contribute to disparities in smoking rate, susceptibility to addiction, and ability to quit smoking. The purpose of this study was to determine the effect of race and gender on the rate of nicotine metabolism as indicated by the nicotine metabolite ratio (NMR) in adolescent smokers. METHODS: One hundred and fifty-nine adolescent smokers aged 13-17 were given 2mg of deuterium-labeled cotinine (cotinine-d4). The NMR was calculated as the ratio of concentrations of deuterium-labeled 3'-hydroxycotinine (ng/ml) to cotinine-d4 (ng/ml) in saliva and is a validated biomarker of the rate of nicotine metabolism. RESULTS: The sample was 67.3% female and racially mixed. On average, Whites had the fastest rates of metabolism compared with both Blacks/African Americans (p < .01) and Asians (p = .01). The NMR was similar between males and females (p = .70). Among the 19 girls who reported using estrogen-containing contraceptives, there was no significant difference in NMR compared with the 83 girls who did not use contraceptives (p = .24) or the 10 who used progestin-only contraceptives (p = .45). CONCLUSIONS: Among adolescent smokers, racial variations in rates of nicotine metabolism were similar to those that have been reported in adult smokers. In contrast to findings in adult smokers, the NMR did not vary significantly by gender or self-reported hormone use.

A multi-route model of nicotine-cotinine pharmacokinetics, pharmacodynamics and brain nicotinic acetylcholine receptor binding in humans.

The pharmacokinetics of nicotine, the pharmacologically active alkaloid in tobacco responsible for addiction, are well characterized in humans. We developed a physiologically based pharmacokinetic/pharmacodynamic model of nicotine pharmacokinetics, brain dosimetry and brain nicotinic acetylcholine receptor (nAChRs) occupancy. A Bayesian framework was applied to optimize model parameters against multiple human data sets. The resulting model was consistent with both calibration and test data sets, but in general underestimated variability. A pharmacodynamic model relating nicotine levels to increases in heart rate as a proxy for the pharmacological effects of nicotine accurately described the nicotine related changes in heart rate and the development and decay of tolerance to nicotine. The PBPK model was utilized to quantitatively capture the combined impact of variation in physiological and metabolic parameters, nicotine availability and smoking compensation on the change in number of cigarettes smoked and toxicant exposure in a population of 10,000 people presented with a reduced toxicant (50%), reduced nicotine (50%) cigarette Across the population, toxicant exposure is reduced in some but not all smokers. Reductions are not in proportion to reductions in toxicant yields, largely due to partial compensation in response to reduced nicotine yields. This framework can be used as a key element of a dosimetry-driven risk assessment strategy for cigarette smoke constituents.

Effect of menthol on nicotine pharmacokinetics in rats after cigarette smoke inhalation.

INTRODUCTION: The effect of menthol on nicotine disposition is important in understanding smoking behaviors among different racial groups. The present study was to evaluate whether menthol affects the pharmacokinetics of nicotine after cigarette smoke inhalation. METHODS: Rats were exposed to mainstream smoke from either a nonmentholated or mentholated cigarette (1 puff/min for 10 min) using a smoke inhalation apparatus. For the multiple-cigarette smoke inhalation, rats received the smoke from either nonmentholated or mentholated cigarette (10 puffs) every 12 hr for a total of 17 cigarettes. Serial blood samples were collected during the 10-min inhalation phase for the single-cigarette smoke or the 17th cigarette inhalation and for 30 hr thereafter. Nicotine and its major metabolite cotinine were assayed by radioimmunoassay methods. RESULTS: Following single-cigarette smoke inhalation, mentholated cigarettes significantly decreased the mean peak concentrations of nicotine in plasma (C(max)) from 27.1 to 9.61 ng/ml and the total area under the plasma concentration-time curves (AUC) from 977 to 391 ng min/ml as compared with those after nonmentholated cigarette smoke inhalation. C(max) and AUC values for cotinine were also significantly reduced by menthol. Similarly after multiple smoke inhalation, C(max), AUC, and the mean average steady-state plasma concentration of nicotine as well as cotinine were significantly lower in mentholated cigarette inhalation. Interestingly, there was a significant increase in the cotinine to nicotine AUC ratio from 13.8 for the nonmentholated to 21.1 for the mentholated cigarette. CONCLUSIONS: These results suggest that menthol in mentholated cigarettes can substantially decrease the absorption and/or increase the clearance of nicotine.

Use of partition coefficients in flow-limited physiologically-based pharmacokinetic modeling.

Permeability-limited two-subcompartment and flow-limited, well-stirred tank tissue compartment models are routinely used in physiologically-based pharmacokinetic modeling. Here, the permeability-limited two-subcompartment model is used to derive a general flow-limited case of a two-subcompartment model with the well-stirred tank being a specific case where tissue fractional blood volume approaches zero. The general flow-limited two-subcompartment model provides a clear distinction between two partition coefficients typically used in PBPK: a biophysical partition coefficient and a well-stirred partition coefficient. Case studies using diazepam and cotinine demonstrate that, when the well-stirred tank is used with a priori predicted biophysical partition coefficients, simulations overestimate or underestimate total organ drug concentration relative to flow-limited two-subcompartment model behavior in tissues with higher fractional blood volumes. However, whole-body simulations show predicted drug concentrations in plasma and lower fractional blood volume tissues are relatively unaffected. These findings point to the importance of accurately determining tissue fractional blood volume for flow-limited PBPK modeling. Simulations using biophysical and well-stirred partition coefficients optimized with flow-limited two-subcompartment and well-stirred models, respectively, lead to nearly identical fits to tissue drug distribution data. Therefore, results of whole-body PBPK modeling with diazepam and cotinine indicate both flow-limited models are appropriate PBPK tissue models as long as the correct partition coefficient is used: the biophysical partition coefficient is for use with two-subcompartment models and the well-stirred partition coefficient is for use with the well-stirred tank model.

Hair biomarkers as measures of maternal tobacco smoke exposure and predictors of fetal growth.

INTRODUCTION: Most biomarker studies of the effects of maternal smoking on fetal growth have been based on a single blood or urinary cotinine value, which is inadequate in capturing maternal tobacco exposure over the entire pregnancy. We used hair biomarkers to compare the associations of maternal self-reported smoking, hair nicotine, and hair cotinine with birth weight for gestational age (BW for GA) among active and passive smokers during pregnancy. METHODS: We collected maternal hair in the immediate postpartum period and measured nicotine and cotinine concentrations averaged over the pregnancy in 444 term controls drawn from 5,337 participants in a multicenter nested case-control study of preterm birth. BW for GA Z-score and small for gestational age (SGA) were based on Canadian population-based standards. RESULTS: The addition of hair nicotine to multiple linear regression models containing self-reported active smoking, hair cotinine, or both explained significantly more variance in the BW for GA Z-score (p = .01, .03 and .04, respectively). Similarly, women with hair nicotine, but not cotinine, at or above the median value had a significant increase in the risk of SGA birth (odds ratio: 3.07, 95% CI: 1.25-7.52). No significant association was observed between maternal passive smoking and BW for GA based on hair biomarkers. CONCLUSIONS: Hair nicotine is a better predictor of reductions in BW for GA than either hair cotinine or self-report. Our negative results for passive smoking suggest that previously reported small but significant effects may be explained by misclassification of active smokers as passive smokers based on self-report.

Role of CYP2A5 in the clearance of nicotine and cotinine: insights from studies on a Cyp2a5-null mouse model.

CYP2A5, a mouse cytochrome P450 monooxygenase that shows high similarities to human CYP2A6 and CYP2A13 in protein sequence and substrate specificity, is expressed in multiple tissues, including the liver, kidney, lung, and nasal mucosa. Heterologously expressed CYP2A5 is active in the metabolism of both endogenous substrates, such as testosterone, and xenobiotic compounds, such as nicotine and cotinine. To determine the biological and pharmacological functions of CYP2A5 in vivo, we have generated a Cyp2a5-null mouse. Homozygous Cyp2a5-null mice are viable and fertile; they show no evidence of embryonic lethality or developmental deficits; and they have normal circulating levels of testosterone and progesterone. The Cyp2a5-null mouse and wild-type mouse were then used for determination of the roles of CYP2A5 in the metabolism of nicotine and its major circulating metabolite, cotinine. The results indicated that the Cyp2a5-null mouse has lower hepatic nicotine 5'-hydroxylation activity in vitro, and slower systemic clearance of both nicotine and cotinine in vivo. For both compounds, a substantially longer plasma half-life and a greater area under the concentration-time curve were observed for the Cyp2a5-null mice, compared with wild-type mice. Further pharmacokinetics analysis confirmed that the brain levels of nicotine and cotinine are also influenced by the Cyp2a5 deletion. These findings provide direct evidence that CYP2A5 is the major nicotine and cotinine oxidase in mouse liver. The Cyp2a5-null mouse will be valuable for in vivo studies on the role of CYP2A5 in drug metabolism and chemical toxicity, and for future production of CYP2A6- and CYP2A13-humanized mouse models.

A comparison of exposure to carcinogens among roll-your-own and factory-made cigarette smokers.

Consumption of roll-your-own (RYO) tobacco is rising, but little is known about its in vivo delivery of toxins relative to factory-made (FM) cigarettes. To start to address this issue, this study compared the concentrations of metabolites of recognized human carcinogens in smokers of RYO tobacco and FM cigarettes. We opportunistically recruited 127 FM and 28 RYO cigarette smokers in central London and collected saliva and urine samples. Saliva samples were assayed for cotinine while urinary samples were assayed for 1-hydroxypyrene (1-HOP) and total 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), metabolic markers of polycyclic aromatic hydrocarbons and tobacco-specific N-nitrosamines, respectively. Data on socio-demographic, anthropometric and puffing characteristics were also obtained. Both unadjusted and adjusted analyses (controlling for age, sex, body mass index, puff flow, puff duration and cotinine) showed no difference in metabolic markers between RYO and FM cigarette smokers. However, significant main effects for cotinine levels and sex were observed in adjusted analyses. Greater levels of cotinine were associated with a greater concentration of both 1-HOP (B = 0.002, P = 0.037) and NNAL (B = 0.002, P < 0.001). In addition, women had significantly greater concentrations of urinary 1-HOP (B = 0.679, P = 0.004) and total NNAL metabolites (B = 0.117, P = 0.024) than men, irrespective of the type of cigarettes smoked. More research is now needed to confirm these findings and gender-specific effects in a larger, representative sample. However, results do not support the common belief that RYO cigarettes are less harmful than manufactured cigarettes.

Glucuronidation of nicotine and cotinine by UGT2B10: loss of function by the UGT2B10 Codon 67 (Asp>Tyr) polymorphism.

Nicotine, the major addicting agent in tobacco and tobacco smoke, undergoes a complex metabolic pathway, with approximately 22% of nicotine urinary metabolites in the form of phase II N-glucuronidated compounds. Recent studies have shown that UGT2B10 is a major enzyme involved in the N-glucuronidation of several tobacco-specific nitrosamines. In the present study, microsomes of UGT2B10-overexpressing HEK293 cells exhibited high N-glucuronidation activity against both nicotine and cotinine with apparent KM's that were 37- and 3-fold lower than that observed for microsomes of UGT1A4-overexpressing cells against nicotine and cotinine, respectively. The KM of microsomes from wild-type (WT) UGT2B10-overexpressing cells for nicotine and cotinine was similar to that observed for human liver microsomes (HLM) against both substrates. The level of glucuronidated nicotine or cotinine in 112 HLM samples was correlated with UGT2B10 genotype; the levels of nicotine- and cotinine-glucuronide were 21% to 30% lower in specimens from subjects with the UGT2B10 (*1/*2) genotype compared with specimens from subjects with the WT UGT2B10 (*1/*1) genotype; a 5- and 16-fold lower level of nicotine- and cotinine-glucuronide formation, respectively, was observed in HLM from subjects with the UGT2B10 (*2/*2) genotype. In contrast to the relatively high activity observed for cells overexpressing WT UGT2B10 in vitro, little or no glucuronidation was observed for microsomes from cells overexpressing the UGT2B10*2 variant against either nicotine or cotinine. These data suggest that UGT2B10 is the major hepatic enzyme involved in nicotine/cotinine glucuronidation and that the UGT2B10*2 variant significantly reduces nicotine- and cotinine-N-glucuronidation formation and plays an important role in nicotine metabolism and elimination.

Genome-wide linkage of cotinine pharmacokinetics suggests candidate regions on chromosomes 9 and 11.

Characterizing cotinine pharmacokinetics is a useful way to study nicotine metabolism because the same liver enzyme is primarily responsible for the metabolism of both, and the clearances of nicotine and cotinine are highly correlated. We conducted a whole-genome linkage analysis to search for candidate regions influencing quantitative variation in cotinine pharmacokinetics in a large-scale pharmacokinetic study with 61 families containing 224 healthy adult participants. The strongest linkage signal was identified at 135 cM of chromosome 9 with LOD = 2.81 and P = 0.0002; two other suggestive linkage peaks appear at 31.4 and 73.5 cM of chromosome 11 with LOD = 1.96 (P = 0.0013) and 1.94 (P = 0.0014). The confidence level of the linkage between the three genome regions and cotinine pharmacokinetics is statistically significant with a genome-wide empirical probability of P = 0.029.

Relationship between skin melanin index and nicotine pharmacokinetics in African American smokers.

BACKGROUND: Blacks bear a disproportionate burden of smoking-related diseases and experience greater difficulty quitting smoking than Whites. Nicotine has a high affinity for melanin, and it has been hypothesized that melanin levels might influence nicotine pharmacokinetics and enhance dependence. The aim of this study was to evaluate the hypothesis that melanin affects nicotine disposition kinetics in humans. METHODS: Forty-four Black participants were administered intravenous infusions of deuterium-labeled nicotine and cotinine. Plasma concentrations of nicotine and cotinine were measured, and pharmacokinetic parameters were estimated. The constitutive and facultative melanin indexes were measured using a dermaspectrophotometer. RESULTS: The median constitutive melanin index was 60.7 (32.8-134.7) and the median facultative melanin index 68.1 (38.6-127.1). The mean (±SD) nicotine elimination half-life was 136 min (±33.5), clearance was 1237 mL/min (±331), and Vss was 204 L (±66), or 2.6 L/kg (±0.7). No evidence of significant differences was found in nicotine pharmacokinetic parameters by comparing participants in different melanin index quartiles (outliers with very high melanin index had similar pharmacokinetic values to others). Differences were not statistically significant when adjusted for age, BMI, sex and CYP2A6 genotype or the nicotine metabolite ratio (NMR), and no evidence of significant correlations were found between melanin (facultative or constitutive) and the pharmacokinetic parameters of nicotine or cotinine or tobacco dependence measures. CONCLUSIONS: Based on our finding in this group of Black smokers, we could not confirm the hypothesis that melanin significantly affects nicotine disposition kinetics or measures of tobacco dependence.

An anti-EGFR × cotinine bispecific antibody complexed with cotinine-conjugated duocarmycin inhibits growth of EGFR-positive cancer cells with KRAS mutations.

Antibody-drug conjugates (ADCs) can selectively deliver cytotoxic agents to tumor cells and are frequently more potent than naked antibodies. However, optimization of the conjugation process between antibodies and cytotoxic agents and characterization of ADCs are laborious and time-consuming processes. Here, we describe a novel ADC platform using a tetravalent bispecific antibody that simultaneously binds to the tumor-associated antigen and a hapten conjugated to a cytotoxic agent. We selected cotinine as the hapten because it is not present in biological systems and is inert and nontoxic. We prepared an anti-epidermal growth factor receptor (EGFR) × cotinine bispecific antibody and mixed it with an equimolar amount of cotinine-conjugated duocarmycin to form the ADC. This ADC showed significant in vitro and in vivo antitumor activity against EGFR-positive, cetuximab-refractory lung adenocarcinoma cells with KRAS mutations.

Nicotine metabolism and CYP2A6 activity in a population of black African descent: impact of gender and light smoking.

Genetic variation in CYP2A6 (the main nicotine metabolizing enzyme) accounts for some, but not all, of the interindividual and interethnic variability in the rates of nicotine metabolism. We conducted a nicotine kinetic study in smokers and nonsmokers of black African descent (N=190), excluding those with common genetic variants in CYP2A6, to investigate the association of demographic variables with CYP2A6 activity (3HC/COT ratio) and nicotine disposition kinetics (estimated nicotine AUC). An additional aim was to examine whether impaired CYP2A6 activity and/or nicotine disposition kinetics were associated with lower cigarette consumption in a population of light smokers (mean

Respiratory retention of nicotine and urinary excretion of nicotine and its five major metabolites in adult male smokers.

Urinary excretion of nicotine and its five major metabolites (nicotine-N-glucuronide, cotinine, cotinine-N-glucuronide, trans-3'-hydroxycotinine, and trans-3'-hydroxycotinine-O-glucuronide), expressed as nicotine equivalents (NE), has been used as a biomarker of smoking-related nicotine exposure. In this open-label, single center study, we investigated the relationship between nicotine retention from smoking and urinary excretion of NE in adult smokers. After a 4-day washout period, 16 adult male smokers smoked 6 cigarettes per day for four consecutive days according to three predefined smoking patterns: no inhalation (Pattern A), normal inhalation (Pattern B), and deep inhalation (Pattern C). The amount of nicotine retained in the respiratory tract during smoking was estimated from the difference between the amounts of nicotine delivered and exhaled. The daily excretion of urinary NE was measured in 24h urine samples by LC-MS/MS. The mean (+/-S.D.) amount of nicotine retained was 0.126+/-0.167, 0.960+/-0.214, and 1.070+/-0.223mg/cig for Patterns A, B, and C, respectively. The mean (+/-S.D.) relative retention (the amount retained relative to the amount delivered) was 11.2+/-14.7%, 98.0+/-1.6%, and 99.6+/-0.3% for Patterns A, B, and C, respectively. On the fourth day of smoking, an average of 86+/-20% of the total daily amount of retained nicotine was recovered as NE in 24h urine. Nicotine equivalents was treated as a single component and the data was described by a first-order elimination pharmacokinetic model which assumed instantaneous input and distribution. Based on this model, the elimination half-life of NE was 19.4+/-2.6h, and the NE excretion had reached approximately 96% of the steady state levels by Day 4. Our results suggest that most of the nicotine inhaled from a cigarette is retained (> or =98%) in the lung, and at steady state, daily urine NE excretion reflects approximately 90% of the retained nicotine dose from cigarette smoking.

Biotransformation of caffeine, cotinine, and nicotine in stream sediments: implications for use as wastewater indicators.

Microbially catalyzed cleavage of the imadazole ring of caffeine was observed in stream sediments collected upstream and downstream of municipal wastewater treatment plants (WWTP) in three geographically separate stream systems. Microbial demethylation of the N-methyl component of cotinine and its metabolic precursor, nicotine, also was observed in these sediments. These findings indicate that stream sediment microorganisms are able to substantially alter the chemical structure and thus the analytical signatures of these candidate waste indicator compounds. The potential for in situ biotransformation must be considered if these compounds are employed as markers to identify the sources and track the fate of wastewater compounds in surface-water systems.