Accelerated metabolism of nicotine and cotinine in pregnant smokers

Cigarette smoking is the foremost modifiable risk factor for adverse pregnancy outcomes. Nicotine is a suspected fetal neuroteratogen. There is concern that nicotine may achieve toxic levels during pregnancy if nicotine replacement therapies are prescribed at doses used in the nonpregnant state. Ten healthy, volunteer, pregnant smokers received infusions of deuterium-labeled nicotine and cotinine during pregnancy and again postpartum. From blood and urine measurements, the following were determined: clearance (renal and nonrenal) of nicotine and cotinine, clearance of nicotine via the cotinine pathway (an indicator of CYP2A6 activity), and daily intake of nicotine from smoking. The clearance of nicotine and cotinine was significantly higher (60 and 140%, respectively), and the half-life of cotinine was much shorter (8.8 versus 16.6 h, P < 0.01) during pregnancy. Although plasma levels of cotinine were lower during pregnancy (119 versus 202 ng/ml, P < 0.05), daily intake of nicotine from smoking was similar during pregnancy and postpartum. For a given level of intake, the pharmacologic and toxicologic effects of nicotine during pregnancy are anticipated to be less than expected from nicotine metabolism data in nonpregnant women. Our data indicate that no downward dose adjustment needs to be made for nicotine replacement therapy during pregnancy. Conversely, higher than usual doses of nicotine may be necessary to optimize efficacy. Lower cotinine levels observed during pregnancy do not necessarily reflect less smoke exposure, and cut-off levels used to classify nonsmokers, passive smokers, and active smokers need to be established for pregnancy.     

Ethnic differences in N-glucuronidation of nicotine and cotinine

We previously reported that the metabolism of cotinine, the proximate metabolite of nicotine, is significantly slower in black than in white cigarette smokers. To understand why the metabolism of nicotine and cotinine might differ between blacks and whites, we studied the pattern of nicotine metabolism in blacks and whites. One hundred eight healthy smokers (51 blacks and 57 whites), of similar age, gender distribution, and smoking history, received an i.v. infusion of deuterium-labeled nicotine and cotinine. The clearance of cotinine, the fractional conversion of nicotine to cotinine, and the metabolic clearance of nicotine to cotinine were significantly lower in blacks than in whites. Blacks excreted significantly less nicotine as nicotine-N-glucuronide and less cotinine as cotinine-N-glucuronide than whites, but there was no difference in the excretion of 3'-hydroxycotinine-O-glucuronide. Nicotine and cotinine glucuronidation appeared to be polymorphic, with evidence of slow and fast N-glucuronide formers among blacks but was unimodal with fast conjugators only among whites. Other findings of note included the demonstration of a significant correlation between the distribution volumes of nicotine and cotinine with lean body mass: there was a smaller distribution volume and a shorter half-life for cotinine in women than in men and a smaller volume of distribution of cotinine in blacks than in whites. We conclude that the metabolism of cotinine is slower in blacks than in whites because of both slower oxidative metabolism of nicotine to cotinine (presumably via cytochrome P-450 2A6) and slower N-glucuronidation. Ethnic differences in the metabolism of other drugs undergoing N-glucuronidation should be studied.     

Effects of passive smoking on theophylline clearance

Theophylline disposition was examined in seven passive smokers, defined as nonsmokers with long-term exposure to cigarette smoke, and seven age-matched nonsmokers with minimal smoke exposure. Subjects were given an intravenous infusion of aminophylline (6 mg/kg) and blood samples were drawn before and during the 48-hour postinfusion period. Clearance for passive smokers was 6.01 x 10(-2) L/hr.kg and for nonsmokers, clearance was 4.09 x 10(-2) L/hr.kg (p less than 0.025). Terminal elimination half-life for passive smokers was 6.93 hours versus 8.69 hours for nonsmokers (p less than 0.05). The mean residence time for passive smokers was 9.89 hours. For nonsmokers, the mean residence time was 13.11 hours (p less than 0.05). These measurements were statistically different, whereas there was no difference in volume of distribution between the groups, suggesting that passive smokers metabolize theophylline more rapidly than nonsmokers. Plasma and urine cotinine and nicotine concentrations were measured in all subjects. There was a significant difference between the subject groups in plasma (p less than 0.004) and urine (p less than 0.002) cotinine concentrations. Theophylline clearance correlated with both plasma (r = 0.73, p less than 0.01) and urine (r = 0.79, p less than 0.01) cotinine concentrations. Additional studies should be conducted to further define the pharmacokinetic characteristics of passive smokers and to assess the effects of passive smoking on drugs metabolized by the mixed function oxidase system.     

Effects of cigarette smoking and carbon monoxide on nicotine and cotinine metabolism

OBJECTIVES: To examine the effects of cigarette smoking on the disposition kinetics of nicotine and cotinine, to determine the effects of cigarette smoking on pathways of nicotine and cotinine metabolism, and to test the hypothesis that carbon monoxide inhibits the metabolism of nicotine. STUDY DESIGN: Twelve cigarette smokers were studied in three treatment conditions, each lasting 7 days, during which they smoked cigarettes, breathed carbon monoxide to achieve carboxyhemoglobin levels similar to cigarette smoking, or breathed air. In each treatment condition, subjects received a combined infusion of deuterium-labeled nicotine (d2) and cotinine (d4), with measurement of disposition kinetics and urine metabolite profile. RESULTS: Cigarette smoking significantly inhibited the metabolism of nicotine but had no effect on cotinine metabolism. Cigarette smoking markedly induced the O-glucuronidation of trans-3'-hydroxycotinine but had no effect on the N-glucuronidation of nicotine or cotinine. Carbon monoxide had no effect on nicotine or cotinine kinetics or metabolic profile. CONCLUSIONS: This study confirms previous observations that cigarette smoking inhibits nicotine metabolism but disproves the hypothesis that this effect is due to carbon monoxide. Induction of glucuronidation must be considered in understanding the effects of cigarette smoking on drug metabolism.     

Nicotine metabolite ratio predicts efficacy of transdermal nicotine for smoking cessation

BACKGROUND: Nicotine is metabolized to cotinine, and cotinine is metabolized to 3'-hydroxycotinine (3-HC) by the liver enzyme cytochrome P450 (CYP) 2A6. More rapid metabolism of nicotine may result in lower nicotine blood levels from nicotine replacement products and poorer smoking cessation outcomes. This study evaluated the utility of the 3-HC/cotinine ratio as a predictor of the efficacy of nicotine replacement therapy as an aid for smoking cessation. METHODS: By use of an open-label design, 480 treatment-seeking smokers were randomly assigned to 8 weeks of transdermal nicotine or nicotine nasal spray use, plus behavioral group counseling. Assessments included demographics, smoking history, body mass index, and plasma nicotine, cotinine, and 3-HC concentrations, as well as CYP2A6 genotypes. Smoking cessation was biochemically verified at the end of treatment and at 6-month follow-up. RESULTS: The rate of nicotine metabolism, as indicated by pretreatment 3-HC/cotinine ratio derived from cigarette smoking, predicted the effectiveness of transdermal nicotine at both time points. The odds of abstinence were reduced by almost 30% with each increasing quartile of metabolite ratio (odds ratio, 0.72 [95% confidence interval, 0.57-0.90]; P=.005). Higher metabolite ratios also predicted lower nicotine concentrations (beta=-1.72, t(179)=-3.31, P<.001), as well as more severe cravings for cigarettes after 1 week of treatment (beta=0.32, t(190)=2.91, P=.004). The metabolite ratio did not predict cessation with use of nicotine nasal spray (odds ratio, 1.05 [95% confidence interval, 0.83-1.33]; P=.68). CONCLUSION: The nicotine metabolite ratio might be useful in screening smokers to determine likely success with a standard dose of transdermal nicotine.     

Nicotine, cotinine, and anabasine inhibit aromatase in human trophoblast in vitro.

Epidemiologic studies suggest that women who smoke have lower endogenous estrogen than nonsmokers. To explore the possible link between cigarette smoking and decreased endogenous estrogens, we have examined the effects of constituents of tobacco on estrogen production in human choriocarcinoma cells and term placental microsomes. In choriocarcinoma cell cultures, nicotine, cotinine (a major metabolite of nicotine), and anabasine (a minor component of cigarette tobacco) all inhibited androstenedione conversion to estrogen in a dose-dependent fashion. Removal of nicotine, cotinine, and anabasine from the culture medium resulted in the complete reversal of the inhibition of aromatase. In the choriocarcinoma cell cultures, a supraphysiologic concentration of androstenedione (73 microM) in the culture medium blocked the inhibition of aromatase caused by nicotine, cotinine, and anabasine. In preparations of term placental microsomes, nicotine, cotinine, and anabasine inhibited the conversion of testosterone to estrogen. Kinetic analysis demonstrated the inhibition to be competitive with respect to the substrate. These findings suggest that some nicotinic alkaloids directly inhibit aromatase. This mechanism may explain, in part, the decreased estrogen observed in women who smoke.     

Relationship between smoking status and serum lipids in a hyperlipidemic population and analysis of possible confounding factors.

The aim of our study was to estimate the potential relationship between smoking behavior and other coronary heart disease risk factors in 250 hyperlipidemic patients. We present data obtained through self-reporting of the number of cigarettes smoked per day, measurements of three tobacco markers, and data on dietary habits and lipid variables. We measured cotinine (by HPLC) and thiocyanate and used a recent colorimetric assay for the indirect evaluation of the nicotine metabolites in a single urine specimen. Mean values of nicotine metabolites, expressed as cotinine equivalents, were 6.7, 39.9, and 79.4 mumol/L, respectively, for nonsmokers, light smokers (7.7 cigarettes per day), and heavy smokers (25.8 cigarettes per day). We found that light smokers have higher concentrations of cotinine and nicotine metabolites in proportion to the number of cigarettes smoked per day than do heavy smokers. Thus, the simple colorimetric assay can accurately evaluate smoking status. Hyperlipidemia and smoking are linked by an intricate network of multiple relations. The concentration of high-density lipoprotein (HDL) cholesterol is lower in heavy smokers, and the concentrations of triglycerides and cholesterol are higher. The 0.11 mmol/L difference in HDL cholesterol between light and heavy smokers is close to the results of previous papers; however, when gender, dietary habits (including alcohol intake), and data on body mass index are included in a multiple regression analysis, there is no longer an association between HDL cholesterol concentrations and smoking status. Therefore, these different dietary habits may be confounding factors that partly explain the pattern of lipid variables.     

How the steady-state cotinine concentration in cigarette smokers is directly related to nicotine intake.

The relationship between nicotine intake and steady-state cotinine concentration was studied in a sample of 125 subjects who smoked their usual brands of cigarettes. Nicotine and tar yield of cigarettes was determined with a smoking machine, under standardized conditions. Blood was drawn about 8 hours after the last cigarette was smoked and serum cotinine was measured by high performance liquid chromatography. Cotinine levels ranged from 11 to 400 ng/ml, and nicotine daily intake ranged from 1 to 33 mg/day. Regression analysis and the correlation coefficient, r = 0.919, significant at p less than 0.0001, showed that steady-state cotinine level was linearly and directly related to daily available nicotine, with an increase in correlation coefficient directly related to the increase in tar and nicotine yield. From the findings we also conclude that smokers of low-tar cigarettes do not tend to compensate for lower yields of nicotine.     

Cotinine disposition and effects

Cotinine is the major metabolite of nicotine in man. We studied cotinine disposition kinetics in 28 healthy habitual cigarette smokers. Eight subjects received cotinine fumarate, 4 micrograms base/kg/min IV for 60 min. Mean (+/- SD) metabolic clearance was 60 +/- 12 ml/min and mean renal clearance was 12 +/- 5 ml/min, averaging 17% of total clearance. Steady-state volume of distribution was slightly greater than body weight (mean 88 +/- 17 l). Terminal t 1/2 averaged 15.8 +/- 4.0 hr in these eight subjects and 19.7 +/- 6.5 hr in another 12 subjects who abstained from smoking for 3 days. The effect of urinary acidification and alkalinization on renal clearance of cotinine during cigarette smoking was studied in another group of eight subjects. Compared with baseline (mean urinary pH 5.8, renal clearance 12.3 +/- 5.9 ml/min), renal clearance was increased about 50% by urinary acidification (pH 4.4, clearance 18.6 +/- 10 ml/min), but it was not affected by alkalinization (pH 6.7, clearance 14.0 +/- 10.4 ml/min). Infusion of cotinine to blood concentrations seen in moderately heavy smokers had no effect on heart rate, blood pressure, or skin temperature, measures that are sensitive to effects of nicotine. No spontaneous subjective effects were reported. We conclude that, at levels to which cigarette smokers are generally exposed, cotinine exerts no cardiovascular activity and weak, if any, psychologic activity.     

Simultaneous and sensitive measurement of anabasine, nicotine, and nicotine metabolites in human urine by liquid chromatography-tandem mass spectrometry

BACKGROUND: Determination of nicotine metabolism/pharmacokinetics provides a useful tool for estimating uptake of nicotine and tobacco-related toxicants, for understanding the pharmacologic effects of nicotine and nicotine addiction, and for optimizing nicotine dependency treatment. METHODS: We developed a sensitive method for analysis of nicotine and five major nicotine metabolites, including cotinine, trans-3'-hydroxycotinine, nicotine-N'-oxide, cotinine-N-oxide, and nornicotine, in human urine by liquid chromatography coupled with a TSQ Quantum triple quadrupole tandem mass spectrometer (LC/MS/MS). Urine samples to which deuterium-labeled internal standards had been added were extracted with a simple solid-phase extraction procedure. Anabasine, a minor tobacco alkaloid, was also included. RESULTS: The quantification limits of the method were 0.1-0.2 microg/L, except for nicotine (1 microg/L). Cotinine-N-oxide, trans-3'-hydroxycotinine, nicotine, and anabasine in urine were almost completely recovered by the solid-phase extraction, whereas the mean extraction recoveries of nicotine-N'-oxide, cotinine, and nornicotine were 51.4%, 78.6%, and 78.8%, respectively. This procedure provided a linearity of three to four orders of magnitude for the target analytes: 0.2-400 microg/L for nicotine-N'-oxide, cotinine-N-oxide, and anabasine; 0.2-4000 microg/L for cotinine, nornicotine, and trans-3'-hydroxycotinine; and 1.0-4000 microg/L for nicotine. The overall interday method imprecision and recovery were 2.5-18% and 92-109%, respectively. CONCLUSIONS: This sensitive LC/MS/MS procedure can be used to determine nicotine metabolism profiles of smokers, people during nicotine replacement therapy, and passively exposed nonsmokers. This method avoids the need for a time-consuming and labor-intensive sample enrichment step and thus allows for high-throughput sample preparation and automation.     

The direct barbituric acid assay for nicotine metabolites in urine: a simple colorimetric test for the routine assessment of smoking status and cigarette smoke intake

The qualitative direct barbituric acid (DBA) method of detecting urine nicotine metabolites was modified to make it quantitative. The performance of the quantitative DBA method was compared with the qualitative method and an established cotinine radio-immunoassay (RIA), using a panel of urines from 128 reported smokers and 383 reported non-smokers. The quantitative DBA method results were highly correlated with the cotinine RIA results, r = 0.85. The coefficients of variation for the two methods were 6% and 10%, respectively. Assuming that the reported smoking history was correct the qualitative DBA method gave a smoking detection rate of 91% and a false positive rate of 3%. At cut-off levels chosen to yield the same false positive rate the quantitative DBA method detected 93% of smokers, close to that of 98% detected with the cotinine RIA. The quantitative DBA method can be used to analyse over 170 samples per day compared to about 70 per day by RIA. It is therefore a fast and inexpensive alternative to cotinine assays for the assessment of smoking status and cigarette smoke intake.     

Postoperative opiate analgesia requirements of smokers and nonsmokers

BACKGROUND: Smoking cigarettes and other forms of nicotine administration appear to blunt the perception of pain. Abrupt discontinuation of nicotine in nicotine-dependent patients appears to increase the perception of pain. The clinical importance of nicotine's effect on pain perception is not fully understood. OBJECTIVE: To determine whether smokers who abruptly discontinue smoking as a result of being hospitalized for coronary artery bypass graft (CABG) require more postoperative opiate analgesics than nonsmokers. METHODS: A retrospective review of patients who underwent a CABG was performed. Smokers (n = 20) were compared with nonsmokers (n = 69) with regard to opiate analgesic use during the first 48 hours postoperatively. The use of nonopiate sedatives was also compared between the groups. RESULTS: When normalized for weight and body mass index, smokers required 23% and 33%, respectively, more opiate analgesics than did nonsmokers (p = 0.027 and 0.023, respectively). The percentage of patients who received benzodiazepines postoperatively was similar in the 2 groups. CONCLUSIONS: In this study, smokers deprived of nicotine required a greater amount of opiates in the first 48 hours after CABG than did nonsmokers. Healthcare providers need to be aware of the potential for increased narcotic requirements among nicotine-deprived smokers. Further study is needed to determine whether nicotine replacement lessens the requirement for postoperative analgesics in smokers.     

Facilitated nitration and oxidation of LDL in cigarette smokers

BACKGROUND: Cigarette smoking increases the risk of developing atherosclerosis and ischaemic heart disease. Smoking-induced oxidative stress is considered to favour oxidation of low-density lipoprotein (LDL) and subsequently promotes the atherogenic process. We investigated whether peroxynitrite, a reaction product of cigarette smoke, is involved in facilitated oxidation of LDL in smokers. MATERIALS AND METHODS: Plasma LDL was obtained from 10 healthy asymptomatic cigarette smokers and 10 healthy nonsmokers. The state of enhanced oxidative stress in the plasma was assessed by LDL subfraction assay using anion-exchange high-performance liquid chromatography (AE-HPLC) and measurements of thiobarbituric acid-reactive substances (TBARS), 8-hydroxydeoxyguanosine (8-OHdG), vitamin E, 3-nitrotyrosine and 3-chlorotyrosine. RESULTS: Smokers showed a significantly higher level of TBARS and 8-OHdG as well as a significantly lower level of vitamin E than nonsmokers, even after stopping smoking for 10 h or more. The LDL subfraction assay demonstrated an increase in oxidatively modified LDL, as expressed by lower levels of LDL1 and higher levels of LDL2. The 3-nitrotyrosine levels in apolipoprotein B in LDL were significantly higher in smokers than nonsmokers, while the 3-chlorotyrosine levels remained unchanged. In addition, these changes observed in the smokers were further accelerated within 30 min after resumption of cigarette smoking when compared with the levels before smoking resumption. CONCLUSION: The present study suggests that peroxynitrite plays a significant role in oxidative modification of plasma LDL induced by cigarette smoking.     

Reference interval and subject variation in excretion of urinary metabolites of nicotine from non-smoking healthy subjects in Denmark

BACKGROUND: Passive smoking has been found to be a respiratory health hazard in humans. The present study describes the calculation of a reference interval for urinary nicotine metabolites calculated as cotinine equivalents on the basis of 72 non-smokers exposed to tobacco smoke less than 25% of the day. METHODS: Twenty subjects (passive smokers) exposed to tobacco smoke more than 25% of the day (subjectively assessed) and 32 smokers were used to validate the estimated reference interval. Urine samples were collected three times during the day approximately at 06.30, 17.00 and 22.45 h. RESULTS: Within-subject variation was found to be 89.4, 72.6, and 79.2% and between-subject variation was found to be 64.5, 64.2, and 36.1%. No gender difference could be demonstrated. In general all subjects showed increased concentrations in the afternoon and evening samples compared to the morning samples. Parametric reference interval for excretion of nicotine metabolites in urine from non-smokers was established according to International Union of Pure and Applied Chemistry (IUPAC) and International Federation for Clinical Chemistry (IFCC) for use of risk assessment of exposure to tobacco smoke. The reference interval for urinary cotinine was estimated to be 1.1-90.0 micromol/mol creatinine in morning samples from non-smokers. An intercomparison between the radioimmunoassay (RIA) method used for determination of nicotine metabolites and a gas chromatography-mass spectrometry (GC-MS) method for determination of cotinine was carried out on 27 samples from non-smokers and smokers. Results obtained from the RIA method showed 2.84 [confidence interval (CI): 2.50; 3.18] times higher results compared to the GC-MS method. A linear correlation between the two methods was demonstrated (rho=0.96). CONCLUSION: The RIA method is rapid and adequate for clinical use in the assessment of exposure to tobacco smoke, i.e. ratio between CV(a)/CV(ti) was<0.50.     

Ascorbic acid content and accumulation by alveolar macrophages from cigarette smokers and nonsmokers

The lung is at risk for injury from inhaled oxidants, including components of cigarette smoke; therefore, maintaining a chemical antioxidant defense would be advantageous. The potential for ascorbic acid to assume this protective role was investigated by comparing the total ascorbate content of alveolar macrophages obtained from human smokers and nonsmokers, from hamsters that were exposed to cigarette smoke for 4 to 6 weeks, and from a control group of unexposed hamsters. The abilities of alveolar macrophages from these four sources to accumulate 14C-labeled ascorbic acid and dehydroascorbate were also compared. The total ascorbate content in hamster macrophages was 19.5 +/- 1.7 and 44.3 +/- 2.8 nmol/10(7) cells for nonsmokers and smokers, (n = 5) and 73.8 +/- 13.1 nmol/10(7) cells (n = 13, p less than 0.1) for nonsmokers and smokers, respectively. In both humans and hamsters, the rates of accumulation of ascorbic acid and dehydroascorbate were significantly greater (p less than 0.05) for alveolar macrophages from smokers compared with nonsmokers of the same species. After internalization, greater than or equal to 70% of the dehydroascorbate was reduced to ascorbic acid by alveolar macrophages from nonsmokers and smokers of both species. An aqueous extract of cigarette smoke oxidized significantly more ascorbic acid to dehydroascorbate in vitro than a comparable volume of phosphate-buffered saline solution without smoke. The increased content of total ascorbate in alveolar macrophages from smokers and their enhanced ability to accumulate ascorbic acid and dehydroascorbate in vitro may reflect protective utilization of ascorbic acid under conditions of increased oxidant stress, compared with nonsmokers. In addition, alveolar macrophages may internalize dehydroascorbate that has been generated by oxidants in the alveolar space and reduce it to ascorbic acid so it can be reused as an antioxidant.     

Selective induction of propranolol metabolism by smoking: additional effects on renal clearance of metabolites

The objective of this study was to examine the effect of cigarette smoking on the activities of the three primary pathways governing propranolol metabolism in human subjects, i.e., glucuronidation, side-chain oxidation and ring oxidation. A single 80-mg dose of propranolol p.o. together with 40 muCi of 4-[3H]propranolol was given to six smoking and seven nonsmoking, young, white, male subjects and the oral clearance of propranolol and the partial clearances through these pathways were determined. The oral clearance of propranolol was increased by 77% (P less than .02) in smokers compared with nonsmokers. This apparent induction of propranolol metabolism was due to a 122% increase in side-chain oxidation (P less than .001) and a 55% increase in glucuronidation (P less than .01). There was no significant effect of smoking on aromatic ring oxidation. Smokers had a 30% greater renal clearance of total metabolites (P less than .05), due mainly to a 53% greater renal clearance of the acidic propranolol metabolite naphthoxylactic acid (P less than .001). These data show a selective effect of smoking on propranolol metabolism and suggest that side-chain oxidation and glucuronidation are mediated by isoenzymes inducible by aromatic hydrocarbons. The selective effect of smoking on the renal clearance of the major propranolol metabolite naphthoxylactic acid may be due to induction of a renal transport protein.     

Increased nicotinic receptors in brains from smokers: membrane binding and autoradiography studies.

Chronic administration of nicotine increases the density of neuronal cholinergic nicotinic receptors in cells and in rodent brain, and similar increases have been reported in brains from human smokers. To further examine this phenomenon, we measured nicotinic receptor binding sites in brain regions from matched populations of smokers and nonsmokers. We first measured binding of [3H](+/-)epibatidine ([3H]EB) and [3H]cytisine in homogenate preparations from samples of prefrontal and temporal cerebral cortex. Binding of each radioligand was significantly higher (250-300%) in both cortical regions from brains of smokers. Frozen sections from each of the cerebral cortical regions and the hippocampus were used for autoradiographic analysis of [3H]EB binding. In cerebral cortex, binding was most dense in layer VI in the prefrontal cortex and layers IV and VI in the temporal cortex. Densitometric analysis of [3H]EB binding sites revealed marked increases of 300 to 400% of control in all cortical regions examined from smokers' brains. Binding in the hippocampal formation was heterogeneously distributed, with dense areas of binding sites seen in the parasubiculum, subiculum, and molecular layer of the dentate gyrus, and the lacunosum-moleculare layer of the CA1/2. Binding of [3H]EB was significantly higher in all six regions of the hippocampus examined from brains of smokers compared with nonsmokers. These increases ranged from 160% of control in parasubiculum to 290% in the molecular layer of the dentate gyrus. The increase in nicotinic receptors in the cerebral cortex and hippocampus of smokers may modify the central nervous system effects of nicotine and contribute to an altered response of smokers to nicotine.     

Decreased ceruloplasmin ferroxidase activity in cigarette smokers

Ceruloplasmin is one of the most important antioxidant proteins in serum. Ceruloplasmin functions as a ferroxidase that oxidizes iron to the Fe3+ state, thereby preventing Fe2+-catalyzed lipid peroxidation and cellular damage. Despite increased antigenic amounts of ceruloplasmin, cigarette smoker serum has previously been shown to exhibit significantly less antioxidant activity than non-smoker serum. We demonstrate that the decreased antioxidant activity of cigarette smoker serum may be explained by a decrease in ceruloplasmin ferroxidase activity. Smokers had a 14% decrease in serum ceruloplasmin ferroxidase activity (units per milliliter) compared with nonsmokers. There was a 24% decrease in ferroxidase activity per milligram of ceruloplasmin in smokers compared with nonsmokers (0.32 +/- 0.009 U/mg vs 0.42 +/- 0.020 U/mg, p less than 0.005). Smoker serum also contained significantly less ceruloplasmin-specific antioxidant activity than nonsmoker serum. These observations may explain the decrement in smoker serum antioxidant activity that could predispose cigarette smokers to increased oxidant injury.     

Decreased leukotriene B4 synthesis in smokers' alveolar macrophages in vitro

Recent studies have shown that alveolar macrophages (AM) are able to release leukotrienes (LTs). Since cigarette smoking inhibits the cyclooxygenase pathway of arachidonic acid metabolism in the AM, we evaluated the LT production by AM from smokers and nonsmokers. AM were obtained from 35 volunteers, 16 nonsmokers, and 19 smokers. The cells were incubated under various conditions including stimulation with 30 microM arachidonic acid, 2 microM ionophore A23187, or both. Each experiment was performed in parallel using cells from a smoker and a nonsmoker. Lipoxygenase products were analyzed by reverse-phase high performance liquid chromatography. After stimulation, nonsmokers' AM produced LTB4 and 5-hydroxy-eicosatetraenoic acid (5-HETE). In incubations of AM with arachidonic acid and ionophore, the amounts of products formed were: LTB4, 317 +/- 56 pmol/10(6) cells and 5-HETE, 1,079 +/- 254, mean +/- SEM. No metabolites were generated under control conditions (no stimulation). In all incubations performed, the peptido-LTs (LTC4, LTD4, and LTE4) were undetectable. In comparison with AM from nonsmokers, those from smokers showed a 80-90% reduction of 5-HETE and LTB4 synthesis (P less than 0.05 to P less than 0.001 according to stimulatory conditions). This defective lipoxygenase metabolite production in AM from smokers was observed over a wide range of stimuli concentrations and incubation times; AM from smokers also had lower levels of intracellular (esterified) 5-HETE than nonsmokers' AM. We also studied blood polymorphonuclear leukocytes (PMNL) and no difference in the synthesis of 5-lipoxygenase products in these cells was noticed between smokers and nonsmokers. These data show that cigarette smoking causes a profound inhibition of the 5-lipoxygenase pathway in AM but not in blood PMNL.     

Nicotine metabolite ratio as an index of cytochrome P450 2A6 metabolic activity

BACKGROUND: Nicotine and a variety of other drugs and toxins are metabolized by cytochrome P450 (CYP) 2A6. Our objective was to evaluate the use of oral nicotine with measurement of the trans-3'-hydroxycotinine (3HC)/cotinine (COT) metabolite ratio as a noninvasive probe of CYP2A6 activity. METHODS: Sixty-two healthy volunteers received an oral solution of deuterium-labeled nicotine (2 mg) and its metabolite cotinine (10 mg). Plasma nicotine and plasma and saliva cotinine and 3HC concentrations were measured over time. RESULTS: The 3HC/COT ratio derived from deuterium-labeled cotinine, measured in either plasma (2-8 hours after administration) or saliva (at 6 hours), was strongly correlated with the oral clearance of nicotine (r = 0.76-0.83, depending on the time of measurement). The 6-hour 3HC/COT ratio from nicotine derived from tobacco in 14 smokers was highly correlated with the ratio derived from deuterium-labeled nicotine (r = 0.88) and was also highly correlated with the oral clearance of nicotine (r = 0.90). Two subjects homozygous for inactive CYP2A6 alleles produced no 3HC, confirming the specificity of the metabolite ratio. The 3HC/COT ratio was also highly correlated with the clearance and half-life of cotinine, consistent with the fact that cotinine is also primarily metabolized by CYP2A6. CONCLUSIONS: The 3HC/COT ratio derived from nicotine either administered as a probe drug or from tobacco use, measured in either plasma or saliva, is highly correlated with the oral clearance of nicotine. The ratio appears to be a useful noninvasive marker of the rate of nicotine metabolism (which is important in studying nicotine addiction and smoking behavior), as well as a general marker of CYP2A6 activity (which is important in studying drug and toxin metabolism).