Epoxide hydrolase activity in human skin.

1 Epoxide hydrolase (EH) activity was measured in biopsied skin (n = 42) using 7-[H3]-styrene oxide as substrate, and separation of the products by high performance liquid chromatography. 2 EH activity (mean +/- s.d.) was present in separated epidermis (139 +/- 105 pmol glycol formed mg-1 min-1) and dermis (165 +/- 120 pmol glycol formed mg-1 microsomal protein min-1). 3 Whole skin EH activity (mean +/- s.d.) varied widely (433 +/- 254 pmol glycol formed mg-1 microsomal protein min-1) 4 No significant difference in EH activity was observed in skin from breast, penis and leg. 5 Skin EH activity does not appear to contribute significantly to the systemic metabolism of epoxide, but may be important in determining the effects of epoxides formed within the epidermis.     

Interindividual variability in the glucuronidation and sulphation of ethinyloestradiol in human liver.

1. Glucuronidation and sulphation of ethinyloestradiol (EE2) was studied in human liver. Microsomal glucuronyltransferase activity was measured in 110 livers whose donors were 71 women and 39 men. Enzyme activity ranged between 12.6 and 242 pmol min-1 mg-1 protein, i.e. over a 19-fold range and the mean (+/- s.d.) glucuronyltransferase activity was 96.8 +/- 47.9 pmol min-1 mg-1 protein. 2. Cytosolic sulphotransferase activity was measured in 138 livers whose donors were 90 women and 48 men. Enzyme activity ranged between 14.4 and 98.2 pmol min-1 mg-1 protein, i.e. over a 7-fold range, and the mean (+/- s.d.) sulphotransferase activity was 43.7 +/- 18.6 pmol min-1 mg-1 protein. 3. Human liver glucuronyltransferase and sulphotransferase activities showed a unimodal distribution pattern. Enzyme activities were neither sex-related nor age-dependent. Sulphotransferase activity did not correlate with glucuronyltransferase activity (n = 80) suggesting that the two enzymes are independently regulated. The ratio of specific glucuronyltransferase to sulphotransferase activity ranged between 0.15 and 8.0 (mean +/- s.d., 2.44 +/- 1.51) and was unimodally distributed.     

Interindividual variability in the N-sulphation of desipramine in human liver and platelets.

1. The activity of N-sulphotransferase (N-ST) with desipramine (DMI) as substrate was measured in 118 human liver specimens, in platelets obtained from 105 subjects, in 12 specimens of human ileum and colon mucosa and in five specimens of human kidney and lung. 2. N-ST activity ranged between 5.71 and 157 pmol min-1 mg-1 protein in human liver and between 0.27 and 17.3 pmol min-1 mg-1 protein in human platelets. 3. Probit analysis was compatible with a unimodal distribution of the data from both liver and platelets. 4. The frequency distribution histograms of N-ST were asymmetric, with a positive skew in data from both liver and platelets. The mode, median and mean of N-ST were 16.4, 30.2 and 40.4 pmol min-1 mg-1 protein in liver, and 2.12, 3.61 and 3.82 pmol min-1 mg-1 protein in platelets, respectively. After logarithmic transformation of N-ST activity, the frequency distribution histogram was symmetric for data from both liver and platelets. 5. In extrahepatic tissues, the average (+/- s.d.) N-ST activity (pmol min-1 mg-1 protein) was 22.2 +/- 22.8 (ileum), 20.9 +/- 26.9 (colon), 12.4 +/- 5.5 (renal cortex), 9.3 +/- 2.8 (renal medulla) and 4.2 +/- 1.1 (lung). N-ST is widely distributed in the body and the intestine is the extrahepatic tissue with the highest N-ST activity.     

Polymorphic hydroxylation of perhexiline in vitro

AIMS: The aims of this study were to examine the in vitro enzyme kinetics and CYP isoform selectivity of perhexiline monohydroxylation using human liver microsomes. METHODS: Conversion of rac-perhexiline to monohydroxyperhexiline by human liver microsomes was assessed using a high-performance liquid chromatography assay with precolumn derivatization to measure the formation rate of the product. Isoform selective inhibitors were used to define the CYP isoform profile of perhexiline monohydroxylation. RESULTS: The rate of perhexiline monohydroxylation with microsomes from 20 livers varied 50-fold. The activity in 18 phenotypic perhexiline extensive metabolizer (PEM) livers varied about five-fold. The apparent Km was 3.3 +/- 1.5 micro m, the Vmax was 9.1 +/- 3.1 pmol min-1 mg-1 microsomal protein and the in vitro intrinsic clearance (Vmax/Km) was 2.9 +/- 0.5 micro l min-1 mg-1 microsomal protein in the extensive metabolizer livers. The corresponding values in the poor metabolizer livers were: apparent Km 124 +/- 141 micro m; Vmax 1.4 +/- 0.6 pmol min-1 mg-1 microsomal protein; and intrinsic clearance 0.026 micro l min-1 mg-1 microsomal protein. Quinidine almost completely inhibited perhexiline monohydroxylation activity, but inhibitors selective for other CYP isoforms had little effect. CONCLUSIONS: Perhexiline monohydroxylation is almost exclusively catalysed by CYP2D6 with activities being about 100-fold lower in CYP2D6 poor metabolizers than in extensive metabolizers. The in vitro data predict the in vivo saturable metabolism and pharmacogenetics of perhexiline.     

Age-related changes in the activities of epoxide hydrolases in different tissues of mice

Age-related changes in epoxide hydrolase (EH) activity in the liver, kidney, lung, and intestine were studied in male C57BL/6 mice of 1 through 30 months of age. Hepatic cytosolic EH activity increased until 15 months after which there was a decline of 59% during senescence (30 months). Hepatic EH activity in the mitochondrial fraction increased until 4 months and decreased thereafter with a 43% decline by 30 months. The hepatic microsomal EH activity increased until 6 months followed by decline of 32% by 30 months. All of these increases and declines were statistically significant. Renal cytosolic EH showed maximum activity at 6 months after which the activity decreased significantly with age. However, renal EH activity in the mitochondrial fraction, in general, did not change substantially with age. Although changes in renal microsomal EH activity were small, the decrease in activity at 9-18 months was significantly lower than at 1 month and 23-30 months. EH activity in the cytosolic, mitochondrial and microsomal fractions of kidney was 2.5-, 2-, and 10-fold less, respectively, than that found in similar subcellular fractions of liver. Cytosolic EH activity in lung and intestine and lung microsomal EH showed variations with age. The intestinal microsomal EH was not detectable under the experimental conditions used.     

The effect of alcoholic cirrhosis on the activities of microsomal aldrin epoxidase, 7-ethoxycoumarin O-de-ethylase and epoxide hydrolase, and on the concentrations of reduced glutathione in human liver

Activities of the microsomal mono-oxygenases 7-ethoxycoumarin O-de-ethylase (EOC) and aldrin epoxidase (AE), together with microsomal epoxide hydrolase (EH) activity and concentrations of reduced glutathione (GSH) have been measured in liver from patients with alcoholic cirrhosis and in normals. Activities of both mono-oxygenases were significantly reduced in alcoholic cirrhosis. EOC activity (pmol 7-OH coumarin formed/mg microsomal protein/min) was 108.0 +/- 10.6 (n = 8) in normals and 60.9 +/- 11.6 (n = 8) in alcoholic cirrhosis (P less than 0.01). AE activity (pmol dieldrin formed/mg microsomal protein/min) was 58.9 +/- 9.5 (n = 11) in normal liver biopsies and 29.9 +/- 8.6 (n = 9) in alcoholic cirrhosis (P less than 0.05). Microsomal EH activity (nmol styrene glycol formed/mg microsomal protein/min) was similar in normals (39.2 +/- 4.4, n = 11) and alcoholic cirrhosis (40.5 +/- 9.1, n = 6). GSH concentrations (microgram GSH/g liver tissue) were lower (P less than 0.01) in alcoholic cirrhosis (792 +/- 73, n = 10) compared to normals (1182 +/- 76, n = 6).     

Nicotine dependence, symptoms and oxidative stress in male patients with schizophrenia.

The high rate of smoking in schizophrenia may reflect patients' attempts to reduce the side effects of antipsychotic medications, and one mechanism for this reduction may be a reduction in oxidative stress and free radical-mediated brain damage that may contribute to schizophrenic symptoms and to complications of its treatment. Symptoms were assessed with the Positive and Negative Syndrome Scale (PANSS), side effects were assessed with the Simpson and Angus Rating Scale (SAS), and malondialdehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and catalase (CAT) activities were measured in plasma. All of these measures were compared in 130 male inpatients with DSM-IV schizophrenia: 104 smokers and 26 non-smokers. The results showed that the positive PANSS symptoms were lower in smokers than non-smokers (14.5 vs 17.5), while the negative symptoms were lower in those who smoked more cigarettes (r=-0.23). The SAS showed no differences. The CAT activity was correlated with both GSH-Px and SOD activities. Of the three enzymes only the CAT activity was significantly higher in smokers than non-smokers (2.9 vs 1.6 U/ml), but greater SOD activity correlated more cigarettes smoked (r=0.24). Consistent with some protection against oxidative stress, MDA also was significantly lower in smokers than non-smokers (9.2 vs 14.4 nmol/ml). The fewer positive symptoms in smokers and fewer negative symptoms in those who smoked more cigarettes may be a selection bias, but appears to be associated with decreased oxidative stress and lipid peroxidation in schizophrenics who smoke tobacco.     

Usefulness of salivary trans-3'-hydroxycotinine concentration and trans-3'-hydroxycotinine/cotinine ratio as biomarkers of cigarette smoke in pregnant women

Nicotine is rapidly and extensively metabolized in humans and negatively impacts the developing fetus. The concentrations of nicotine, cotinine, trans-3'-hydroxycotinine (hydroxycotinine), and norcotinine in pregnant smokers' oral fluid were evaluated to determine usefulness as biomarkers of cigarette smoking. Sixteen participants were divided into two groups: eight light smokers (LS) who smoked < or =10 cigarettes/day and eight heavy smokers (HS) who smoked > or =20 cigarettes/day. Oral fluid specimens (n=415) were collected throughout pregnancy and analyzed with solid-phase extraction followed by gas chromatography-mass spectrometry-electron impact selected ion monitoring. Median concentrations of nicotine, cotinine, and hydroxycotinine in oral fluid of LS ranged from 241.1 to 622.0, 80.6 to 387.5, and 14.4 to 117.7 ng/mL and for HS 146.5-1372.2, 66.0-245.8, and 38.3-184.4 ng/mL, respectively. Salivary cotinine and hydroxycotinine concentrations were significantly correlated in LS (r = 0.55, p < 0.01) and HS (r = 0.74, p < 0.01). Ratios of hydroxycotinine/cotinine in oral fluid from pregnant women averaged 0.30 +/- 0.18 (range, 0.07-1.05) for LS and 0.68 +/- 0.25 (range, 0.29-1.83) for HS. Based on these preliminary data, the best ratio to differentiate light from heavy pregnant smokers was 0.41. Salivary hydroxycotinine and cotinine concentrations are both good biomarkers of cigarette smoking. Determining the hydroxycotinine/cotinine ratio may differentiate light from heavy tobacco use and help predict increased fetal tobacco exposure.     

Effects of low-yield-cigarette smoke inhalation on rat lung macrophages

It has been suggested that low-yield cigarettes (LYC) may be less hazardous and that smokers of these cigarettes are exposed to less tar, nicotine, and carbon monoxide. Recent studies have challenged this and question the analysis techniques for measuring yields of these cigarettes. Because published LYC contents may not reflect tissue toxicity and because compensatory puffing behaviors may alter smoke delivery to end-organ tissues, we studied the effect of smoke from a typical LYC on phagolysosome fusion and phagocytosis in alveolar macrophages of rats that chronically inhaled the smoke generated by an intermittently puffing apparatus. Alveolar macrophages were obtained by lung lavage and established in monolayers. Phagolysosome fusion and phagocytosis were assessed using the acridine orange fluorochrome assay. After 8 wk of exposure, there was no difference in phagolysosome fusion between controls and smokers. Carboxyhemoglobin levels in controls versus smokers were 1.36 +/- 0.09% versus 2.13 +/- 0.32% (mean +/- SE) (p = 0.06). A group of animals was similarly exposed, but the side pores of the cigarette filters were sealed with tape to simulate the compensatory behaviors often used by LYC smokers of occluding filter pores with their lips or fingertips. This significantly increased smoke exposure, and the carboxyhemoglobin level of the smokers increased to 7.0 +/- 1.4% (versus controls, p less than or equal to 0.01). Cells from these rats showed alterations in phagocytosis and in phagolysosome fusion compared with alveolar macrophages of control rats. These data suggest that the tobacco in LYC may have toxic effects similar to those of high-yield cigarettes and that LYC are likely to be less hazardous only if smoked in a fashion similar to that of a smoke-generating apparatus.     

Cyclosporin A treatment enhances angiotensin converting enzyme activity in lung and serum of rats

Nephrotoxicity and arterial hypertension are the most common side effects of treatment with cyclosporin A (CSA). Its effects on angiotensin converting enzyme (ACE) activity in the renal cortex, lung and serum of nephrotoxic rats have been investigated. Wistar rats were treated with CSA (20 mg kg-1 day-1 i.p.) or vehicle (olive oil containing 10% ethanol) for 14 days. On day 15, the rats were killed and ACE activity determined by radiometric assay using [3H]hippuryl-glycyl-glycine as substrate. CSA treatment resulted in a decrease in creatinine clearance, urine flow and body weight and a significant increase in serum and lung ACE activities (436 +/- 9 vs 391 +/- 7 nmol mL-1 min-1, P less than 0.001; 184 +/- 8 vs 142 +/- 10 nmol mg-1 min-1 P less than 0.01, respectively). In contrast, renal cortex ACE activity was reduced in the CSA-treated rats (0.35 +/- 0.02 vs 0.51 +/- 0.02 nmol mg-1 min-1, P less than 0.01). ACE activities in the renal cortex and serum were not affected by treatment with gentamicin (80 mg kg-1 day-1) for 11 days. In rats treated simultaneously with CSA and captopril (50 mg kg-1 day-1) ACE activity in the serum, lung and renal cortex was inhibited by 95, 93 and 92%, respectively. These changes in ACE activity were associated with a decreased systolic blood pressure in the rats receiving CSA and captopril. Therefore, ACE activity in the serum and lung of CSA-treated rats was increased, while its activity in the renal cortex was reduced.(ABSTRACT TRUNCATED AT 250 WORDS)     

The smoking habits of New Zealand doctors: a review after ten years.

A second survey of New Zealand doctors' smoking habits in 1972 elicited an 83 percent response from 3113 doctors. 38.5 percent had never smoked compared with 23.8 percent in a 1963 survey; 29.2 had given up smoking, and 33.3 percent still smoked. Only 21.3 percent smoked cigarettes compared with 35.3 percent in 1963. There has been a sustantial increase in non-smokers among recent graduates. Both sexes now smoke cigarettes less frequently but pipe and cigar smoking by male doctors has risen sharply. Obstetricians smoke cigarettes more often than other groups of doctors, while pathologists, medical administrators and academics smoke the least. Giving up smoking was not difficult for most former smokers except for the heavy smokers who now make up most of the persistent smoker group.     

Transfer of dideoxyinosine across the human isolated placenta.

1. The metabolism of imipramine (N-demethylation and 2-hydroxylation) was studied in relation to the activity of S-mephenytoin 4'-hydroxylase in human liver microsomes. 2. Eadie-Hofstee plots for the formation of despiramine and 2-hydroxyimipramine were biphasic, suggesting that at least two enzymes are involved in both the N-demethylation and 2-hydroxylation of imipramine by human liver microsomes. 3. The respective mean (+/- s.d.) kinetic parameters for the N-demethylation and 2-hydroxylation of imipramine derived from a two-enzyme kinetic analysis were: Km1 = 1.1 +/- 0.4 and 1.6 +/- 0.6 microM, Vmax1 = 0.11 +/- 0.03 and 0.15 +/- 0.07 nmol mg-1 min-1, and Vmax1/Km1 = 0.10 +/- 0.02 and 0.09 +/- 0.04 ml mg-1 min-1; Km2 = 214 +/- 84 and 257 +/- 148 microM, Vmax2 = 2.22 +/- 0.69 and 0.53 +/- 0.15 nmol mg-1 min-1, and Vmax2/Km2 = 0.011 +/- 0.001 and 0.003 +/- 0.002 ml mg-1 min-1. 4. With regard to imipramine N-demethylation and 2-hydroxylation at 2 microM (representing high-affinity reactions) and at 400 microM (representing low-affinity reactions), only N-demethylation at 2 microM showed a close correlation with the 4'-hydroxylation of S-mephenytoin (rs = 0.952, P < 0.01; n = 10 livers). 5. Concentrations up to 250 microM S-mephenytoin inhibited the N-demethylation of imipramine (2 microM), but no further inhibition was observed using concentrations from 250 to 750 microM. 6. Imipramine inhibited S-mephenytoin 4'-hydroxylation competitively with a Ki value of 12.5 microM.(ABSTRACT TRUNCATED AT 250 WORDS)     

Shorter time to first cigarette of the day in menthol adolescent cigarette smokers

Menthol smoking is thought to contribute to the addictiveness of smoking. Given the high prevalence of menthol smoking among youth, the aim of the current analysis was to examine differences in consumption and tobacco dependence, including smoking urgency among menthol and non-menthol adolescent smokers. Data for the current analysis were collected from telephone interviews with adolescent smokers applying to a cessation treatment study. Of 572 adolescent smokers (mean age=15.6+/-1.6 years; 55.1% female; 46.9% African American, 48.2% European American), 531 smoked menthol cigarettes and 41 smoked non-menthol as their usual brand. Analysis using Fisher's Exact (one-tailed) Test revealed that menthol smokers had a significantly shorter time to first (TTF) cigarette of the day compared to non-menthol smokers (smoking within the first 5 min of the day, 45% vs. 29%, respectively; p<0.04). Independent t tests revealed no significant difference in number of cigarettes per day (CPD) (mean=12.2+/-8.5 vs. 11.4+/-8.8; p<0.28) or Fagerstrom Test for Nicotine Dependence (FTND) scores (3.4+/-1.4 vs. 3.2+/-1.3; p<0.23). While preliminary, our findings suggest greater smoking urgency among menthol compared to non-menthol adolescent cessation-treatment seekers. Further study in a broader sample of adolescent smokers is warranted to elucidate the mechanisms underlying the effects of menthol smoking for youths.     

The increase in urinary excretion of 6 beta-hydroxycortisol as a marker of human hepatic cytochrome P450IIIA induction.

1. Urinary excretion of 6 beta-hydroxycortisol, hepatic microsomal cortisol 6 beta-hydroxylase and the specific content of several forms of cytochrome P450 were measured in 8 to 14 patients before and after treatment with rifampicin (600 mg orally per day for 4 days). 2. Rifampicin treatment produced an average five fold increase in daily excretion of urinary 6 beta-hydroxycortisol. 3. Cortisol 6 beta-hydroxylase activity increased from 15 +/- 6 pmol min-1 mg-1 in organ donors (considered as 'control subjects') to 87 +/- 31 pmol min-1 mg-1 in rifampicin treated patients. 4. Among three forms of human P450 (P450IA, IIC and IIIA), (1), (2), measured by Western blots, only P450IIIA was significantly induced by the antibiotic. 5. Only antibodies against P450IIIA selectively inhibited cortisol 6 beta-hydroxylase in human liver microsomes. 6. Cortisol 6 beta-hydroxylase was correlated with P450IIIA specific content. 7. The urinary level of 6 beta-hydroxycortisol correlated with liver microsomal cortisol 6 beta-hydroxylase and P450IIIA specific content. 8. We conclude that P450IIIA is predominantly responsible for cortisol 6 beta-hydroxylase activity in human liver microsomes and that urinary 6 beta-hydroxycortisol is a marker of the induction of this cytochrome P450.     

Passive smoking exposure of sick children in Hong Kong.

1. This study aims to investigate the extent of passive smoking exposure of sick children in Hong Kong; their father's smoking behaviors and their mother's action to protect the child from environmental tobacco smoke (ETS). 2. This cross-sectional survey was the first phase of a randomized controlled trial on a health education intervention provided by nurses in the general paediatric wards of four major hospitals in Hong Kong. The subjects are non-smoking mothers of sick children admitted to hospital and with a smoking husband living together in the same household with the child. Eligible subjects completed a self-administered baseline questionnaire before entering into the trial. 3. All the 848 children whose mothers completed the questionnaire during December 1997 to April 1998 had a smoking father. They constituted about 24% (95% CI: 22.6-24.9%) of all the sick children during the same period. More than half of the children's fathers (55%; 51.6-58.3%) smoked 1-14 cigarettes per day and about 68% (64.7-71.0%) of them were daily smokers for over 10 years. Over half (53%; 49.4-56.2%) of the fathers smoked 1-14 cigarettes at home every day. About 21% of the fathers smoked near the child occasionally and 31% (27.4-33.6%) smoked 1-14 cigarettes near the child per day. About 16% of the children lived with more than one smoker (2-3). About 86% (83.3-88.0%) of the children had 1-3 smokers who smoked at home and 61% (58.1 - 64.7%) of them had 1-3 smokers who smoked near them every day. However, 70% of the children were reported by their mothers as exposed to ETS at home. This percentage (70%) was less than the percentage (86%) of smokers who smoked at home daily. About 31.1% of the mothers reported symptoms of coughing and running nose (20.6%) in their children when they were exposed to ETS. To protect the child from ETS exposure, 43% of the mothers requested the father not to smoke near the child, 33.1% requested the father to smoke less cigarettes and 31.5% advised the father to quit smoking. 4. In conclusion, sick children in Hong Kong are at risk of exposure to ETS, but not all mothers are aware of the health risks and their actions were inadequate. There is a critical need to promote awareness of ETS exposure and the health risks in the family in Hong Kong so as to reduce illness and hospital admission in children.     

Benzene levels in ambient air and breath of smokers and nonsmokers in urban and pristine environments

Benzene levels in human breath and in ambient air were compared in the urban area of San Francisco (SF) and in a more remote coastal pristine setting of Stinson Beach, Calif. (SB). Benzene analysis was done by gas chromatography-mass spectroscopy (GC-MS). Ambient benzene levels were sevenfold higher in SF (2.6 +/- 1.3 ppb, n = 25) than SB (0.38 +/- 0.39 ppb, n = 21). In SF, benzene in smokers' breath (6.8 +/- 3.0 ppb) was greater than in nonsmokers' breath (2.5 +/- 0.8 ppb) and smokers' ambient air (3.3 +/- 0.8 ppb). In SB the same pattern was observed: benzene in smokers' breath was higher than in nonsmokers' breath and ambient air. Benzene in SF nonsmokers' breath was greater than in SB nonsmokers' breath. Marijuana-only smokers had benzene breath levels between those of smokers and nonsmokers. There was little correlation between benzene in breath and number of cigarettes smoked, or with other benzene exposures such as diet. Of special interest was the finding that benzene in breath of SF nonsmokers (2.5 +/- 0.8 ppb) was greater than that in nonsmokers ambient air (1.4 +/- 0.1 ppb). The same was true in SB, where benzene in nonsmokers breath was greater than ambient air (1.8 +/- 0.2 ppb versus 1.0 +/- 0.1 ppb on d 1 and 1.3 +/- 0.3 ppb versus 0.23 +/- 0.18 ppb on d 2). This suggests an additional source of benzene other than outdoor ambient air.     

Smoking Topography in a Nonlaboratory Environment

The relationships between the number of cigarettes smoked/day and the number of puffs/cigarette, puff duration, and total puff time/cigarette were studied. Data were collected on 12 regular smokers for all cigarettes smoked over a 3-day period in a nonlaboratory environment. Between-subject variability was substantial on each of the topographical measures. Neither the number of cigarettes smoked per day nor the classification of Heavy (> 25 cigarettes/day) vs Moderate (< 25 cigarettes/day) smoking levels was related to the intensity with which cigarettes were smoked. Within-subject consistency on the topography measures indicates that smokers may have relatively unique smoking patterns.

Most studies of smoking in the natural environment employ number of cigarettes/day as their estimate of smoke exposure. However, total smoke exposure is determined by an interaction of various topographical features, including frequency (number of cigarettes/day, number of puffs/cigarette), durational (puff duration, interpuff interval, intercigarette interval), and volumetric (puff volume, inhalation volume) components. Employing cigarettes/day to estimate smoke exposure assumes a consistent relationship between cigarettes/day and other topographical features which contribute to total smoke exposure, but it is not clear that such a relationship exists. Laboratory studies of smoking behavior have found that cigarette frequency may vary independently of these other topographic components of smoking, lichtenstein and Antonuccio (1981) examined smoking topography in 24 male smokers while they smoked a cigarette during two 45-minute sessions. They found that cigarette rate was significantly related to intercigarette interval, but not to puff frequency, puff duration, cigarette duration, and amount of tobacco burned.

Results found in laboratory settings, however, have been found not to necessarily generalize to nonlaboratory environments. For example, OssipKlein, Martin, Lomax, Prue, and Davis (1983) examined six subjects smoking adlib in three settings: natural, clinical, and laboratory. They found that cigarette durations were shorter and that subjects took significantly longer and more puffs in a clinical or laboratory setting compared to a naturalistic setting. Thus, examination of the relationship between topographical features in naturalistic smoking would appear to require direct study outside the laboratory.

The present study is to our knowledge the first to examine topographical features of smoking and the relationship between number of cigarettes smoked/ day and other measures of smoking topography while the subject smoked ad-lib in a nonlaboratory environment. This information would potentially be important in examining the extent of individual differences in smoking topography, in assessing the extent to which cigarettes/day is related to other aspects of smoking behavior, and in determining whether categorizing smokers into smoking groups (e.g., moderate and heavy) on the basis of number of cigarettes/day accurately reflects the amount of total smoke exposure/day.     

Inhibition of human liver microsomal (S)-nicotine oxidation by (-)-menthol and analogues

(-)-Menthol is a widely used flavoring ingredient present in mouthwash, foods, toothpaste, and cigarettes; yet, the pharmacological effects of menthol have not been widely studied. Mentholated cigarette smoking may increase the risk for lung cancer. Many African American smokers smoke mentholated cigarettes, and African Americans have a significantly higher incidence of lung cancer as compared with whites. There may be a relationship between the incidence of lung cancer and the type of cigarette smoked because the use of mentholated cigarettes by white smokers is significantly less and the incidence of lung cancer is less. The mechanism whereby (-)-menthol could increase the health risk of smoking is not known. The results of our in vitro studies herein show that (-)-menthol and synthetic congeners inhibit the microsomal oxidation of nicotine to cotinine and the P450 2A6-mediated 7-hydroxylation of coumarin. Replacement of the alcohol oxygen atom of menthol with other heteroatoms increased the potency of P450 2A6 inhibition. Thus, the K(i) value of (-)-menthol for inhibition of microsomal nicotine oxidation was 69.7 micro M but neomenthyl thiol possesses a K(i) value of 13.8 micro M. Menthylamine inhibited nicotine oxidation with a K(i) value of 49.8 micro M, but its hydroxylamine derivative gave an IC(50) value of 2.2 micro M. A series of 16 menthol derivatives and putative metabolites were procured or chemically synthesized and tested as inhibitors of P450 2A6. While highly potent inhibition of P450 2A6 was not observed for the menthol analogues examined, it is nevertheless possible that smoking mentholated cigarettes leads to inhibition of nicotine metabolism and allows the smoker to achieve a certain elevated dose of nicotine each day. This may be another example of self-medication to obtain the desired effect of nicotine.     

The effect of cigarette burn time on exposure to nicotine and carbon monoxide in adult smokers

Cigarette burn time (CBT), conventionally defined as the time a cigarette burns during smoking, can be affected by cigarette design and smoking behavior. A previous study showed a strong negative correlation between CBT and nicotine yield under machine smoking conditions. This study for the first time examined the relationship of CBT and exposure to nicotine and carbon monoxide in adult smokers in a controlled clinical study. 24h nicotine equivalents excretion (NE), carboxyhemoglobin (COHb) and CBT were measured in two groups of 20 adults smoking Marlboro Lights and 20 adults smoking Marlboro Ultra on two consecutive days. Approximately 20% of the total variability in CBT was attributed to cigarette brand, 34% to smokers and 1% to study day. The exposure index, defined as the number of cigarettes smoked per day divided by average daily CBT for each smoker, accounted for a large proportion of the total variability in NE (R(2)=0.79-0.91) and COHb (R(2)=0.85-0.90). We conclude that CBT has an important influence on levels of NE and COHb in adult smokers. CBT, along with the number of cigarettes smoked per day, can be used to estimate adult smokers' exposure to nicotine and carbon monoxide.     

Expression of cytochromes P450 1A1 and 1B1 in human lung from smokers, non-smokers, and ex-smokers

Cytochromes P450 1A1 and 1B1 are known to bioactivate procarcinogens such as polycyclic aromatic hydrocarbons (PAHs) found in cigarette smoke and are inducible via an Ah receptor-mediated mechanism. The aim of this study was to examine the levels of expression of CYP1A1 and CYP1B1 in samples of lung from smokers (n = 18), non-smokers (n = 7), and ex-smokers (n = 7). Using immunoglobulin preparations of highly specific polyclonal antibodies and immunoblot analysis of microsomes from lung tissues, we determined the specific content for CYP1A1 and CYP1B1. For CYP1A1, we found median expression levels of 15.5 pmol/mg microsomal protein in smokers, 6.0 pmol/mg microsomal protein in non-smokers, and 19.0 pmol/mg microsomal protein in ex-smokers. The difference in median expression levels of smokers and ex-smokers compared to non-smokers was statistically significant. For CYP1B1, we found median expression levels of 1.8 pmol/mg microsomal protein in smokers, 1.0 pmol/mg microsomal protein in non-smokers, and 4.4 pmol/mg microsomal protein in ex-smokers. The difference in median expression levels between ex-smokers and non-smokers was statistically significant. These results suggest that levels of expression of CYP1A1 and CYP1B1 protein in lung tissues from smokers and ex-smokers are quantitatively greater than in non-smokers. By immunohistochemical analysis, we demonstrated the expression of CYP1A1 and CYP1B1 in normal human alveolar type I and II cells, ciliated columnar epithelial cells lining bronchoalveolar airways, and alveolar macrophages. These results confirm that CYP1A1 is expressed in normal human lung, appears to be induced in smokers, and show interindividual variation; the similar characteristics of CYP1B1 are demonstrated.