Environmental tobacco smoke is one of the most widespread carcinogenic exposures. Accurate measurements of ETS exposure, intake and biological effects are therefore needed considering the substantial number of persons daily exposed and the great amount of scientiic data on its association with chronic diseases. ETS exposure is associated with decreased pulmonary function in humans, especially those with asthma. ETS monitoring studies and the dissemination of the results are very important for educational purposes, especially for young people. In the age group of 15 to 17 years, there are many individuals who have a strong or a very strong dependence on nicotine. As a result, it is necessary to promote smoking cessation and nicotine dependence treatment by recommending pharmaceuticals of substantial nicotine therapy. The only known way to reduce cancer risk in smokers is complete cessation, but many smokers are unable or unwilling to quit. ETS, a class A carcinogen, is considered a preventable occupational health risk. It is the major source of indoor pollution because the estimated exposure of the nonsmoking population to ETS at home or at work is high, 50-75% in the US, for example. Prevention consists of measures to avoid the exposure of non-smokers to ETS: public policies that promote smoke-free environments, restriction of smoking in different areas as the workplace, restaurants, buses, in the vicinity of the pregnant women and children, ventilation, information and education especially for the young population. Nicotine, a major toxic compound of which approximately 95% of our exposure derives from tobacco, is present in the gas phase, indoors, in a concentration of about 30 μg.m-3. It is rapidly metabolised, the half life time being about 2 h Cotinine, one of the nicotine metabolites, with a half life time of 20 h, is the compound most frequently used for assessing tobacco smoke exposure. Cotinine concentration in urine varies from values lower than 1 μg.L-1 to 10 Rg.L-1 in passive smokers, and is higher than 100 lig:L-1 in smokers. Serum cotinine, ng.mL-1, represents a reliable epidemiological marker of nicotine intake and may be helpful in ETS exposure studies. Biomarkers measured in different body luids or tissues including urine, blood, saliva, hair, nails and teeth could differentiate the levels of exposure to tobacco smoke and levels of intake. The inefficiency of methodology to assess ETS exposure, especially in public places, belies the magnitude of its impact, especially in lung cancer. As a consequence, a large number of studies have been dedicated to ETS exposure over the last several years. The methods most used for the ETS monitoring are gas chromatography (GC) with lame ionisation detection (FID) or nitrogen specific detectors, or coupled, with electron impact mass spectrometers (EI-MS) as well as the use of high performance liquid chromatography (HPLC) with UV or MS detectors. This paper presents a gas chromatography-mass spectrometric (GC-MS) quantitative method for nicotine and cotinine levels in the indoor air and in urine, for measuring the levels of airborne nicotine in some public houses and for correlating urinary cotinine and nicotine levels in smoking and non-smoking subjects. Nicotine (98%) was obtained from Merck, Germany. All other reagents were from Comchim (Bucharest, Romania). Urine samples from passive smokers and smokers were collected. Samples were collected from 2 non-smokers, working in 2 different pubs where smoking was allowed, from 1 non-smoker in a laboratory room of 40 m3 and 2 smokers. The non-smoker workers had been ETS exposed for over 6 months. The smokers declared that they smoked more than 10 cigarettes per day. The subjects donated 24-hours urine samples after ETS exposure and eache sample was analysed at once. The air nicotine was measured in the 2 pubs and in the laboratory room, after 4 cigarettes had been smoked. Active charcoal cartridges were used for the adsorption from nicotine from 10 L of air pumped at a flow rate of 400 mL.min-1. The pubs were ventilated but the laboratory was not ventilated during the air sampling. A written consent was obtained from each subject. Nicotine and cotinine were extracted from 10 mL urine in to chloroform/diethyl ether (2 m L.0.5 mL) by liquid-liquid extraction for 1 min. After centrifugation for 3 min, the supernatant was removed and 500 ng of the internal standard (IS), 1,2,3-trichlorobenzene was added to the remaining extract. The extract from the non-smokers urine was concentrated. For analysis 3 μL aliquot samples of the extracts were directly injected into the GC. After passing 10 L air through active charcoal cartridges any adsorbed material was then extracted into dichloromethane for 2 min. After centrifugation, the IS was added and 3 μL aliquot samples were injected into the GC. GC-MS analyses were performed so that the nicotine in air samples and nicotine and cotinine in human urine could be determined simultaneously. A Trace DSQ ThermoFinnigan quadrupole mass spectrometer coupled with a Trace GC was used. Nicotine and cotinine and the IS were separated on a Rtx-5MS capillary column, 15 m x 0.25 mm, 0.25 pm film thickness, using an 11-min temperature programme starting from 50°C, held for 1 min then increased at 20°C•min' to 250°C with the MS in the selected ion monitoring (SIM) mode, or starting from 50°C held for 2 min and then increased at 8°C-min-1 to 310°C, with the MS in the scan mode. In the SIM mode the following important ions from the mass spectra of nicotine, cotinine and 1,2,3-trichlorobenzene were used: m/z 84, 133, 161, 162 for nicotine, m/z 98 and 176 for cotinine and m/z 180 and 182 for the IS. Some caffeine metabolites such as theophylline or theobromine which also have a molecular ion at m/z 180, could also be observed in the urine samples with the temperature programe used. The method was validated for the compounds in the range 0-5 μg.mL-1 and the parameters of the linearity, precision, accuracy and limit of detection were studied. Aliquot samples containing 0.05, 0.1, 0.2, 0.3, 1, 2, 3, 4 and 5 μg.mL-1 nicotine in chloroform were used for the method validation. The same were used in water following the extraction procedure described above. Linearity in the range 0-5 μg.mL-1 gave a regression curve: y = 0.0857x - 0.0728 and a coefficient of regression of 0.99. In the range 0-1 μg.mL-1 the curve was y = 0.0149x + 0.0054 and the regression coefficient of 0.98. The second regression curve was used for small quantities of biomarkers. Table 1 presents the linearity, the limit of detection and the recovery measured. Precision and accuracy were studied on the aliquot sample of 0.3 μg.mL-1. Table 2 presents the results obtained for the sample of 0.3 μg-mL-1 nicotine for precision and accuracy. For the air samples the concentration range studied was 1-10 µg of nicotine. The regression curve obtained by using decane as IS was 0.4325x - 0.054, r = 0.98. Recovery was 10% for the concentration of 10 μg. The use of 1,2,3-trichloromethane as IS improved the limit of detection and improved recovery as shown by the similarity of the values for the urine samples. The high concentration of nicotine in air was measured by using the second regression curve. Table 3 presents nicotine and cotinine measured in air and the concentration in urine. The 2 passive smokers, occupationally exposed to ETS, could be differentiated by the time they were employed under these exposure conditions and the level of nicotine measured. The worker in pub 1 had been employed for more than 6 months but in conditions of lower ETS exposure (>200 cigarettes per day). The worker in pub 2 was much more exposed, but still he had been employed for 6 months (>500 cigarettes per night). In the laboratory, without ventilation, the indoor air nicotine was present at very high concentration near the ash tray and at 3 m distance after 4 cigarettes had been smoked in the room, but ventilation reduced the smoke considerably. The olfactory senses confirmed the reduction found. The GC-MS analysis in the SIM mode of nicotine and the internal standard is shown in Figure 2. The ions selected for the SIM mode nicotine determination are also shown. Urinary nicotine and cotinine detection in a smoker's urine in the scan and SIM mode are shown in Figures 3 and 4. The 2 bio-markers eluted in 8 min. Comparison of the separation chromatograms in the SIM mode, for 1 smoker and 2 non-smokers at different ETS exposure times is observed in Figure 5. The GC-MS method presented for determination of nicotine and cotinine concentrations in urine and air is simple and rapid. The method validation gave the following values: 27% RSD precision and 32% RSD for accuracy. Choosing an internal standard of the same chemical class of compounds or better, a stable isotope labelled nicotine could further improve the validation parameters. The urinary cotinine determination seems to be a more important measure than the air nicotine determination. The time declared for the duration of exposure to nicotine as low, medium or high, or the number of cigarettes smoked indoors, could be sufficient for an estimation of indoor air nicotine. Any association with teeth, hair, blood or saliva cotinine levels could be important especially for ETS exposure studies on children. Although high values of urinary cotinine are found in occupationally exposed passive smokers, >100 µg.L-1, those passive smokers exposed for just a few hours had urinary cotinine levels lower than 0.01 μg.L-1. It takes more than 10h after heavy ETS exposure before higher levels of urinary cotinine are observed in a passive smoker because of the time needed to form the metabolite, cotinine, in urine. A significant correlation between urinary cotinine and nicotine increases and airborne nicotine in passive smokers was observed (r = 0.41 and r = 0.77, respectively). In smokers, the declared number of cigarettes smoked per day and the urinary cotinine showed significant correlation. Further statistical studies on ETS exposure are important for informative and educational purposes. Counselling of mothers and young people should promote smoking cessation, to prevent health risks associated with tobacco smoke.9352
Get your grade
or your money back
using our Essay Writing Service!