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Gea News Settembre 2001 - Danni ai coltivatori di Tabacco in Umbria

Lavoro originale. Fonte: http://brownw.newsreal.com/pages/brownw/Story.nsp?story_id=22816720&ID=brownw&scategory=Smoking+%26+Health


Systemic nicotine exposure in tobacco harvesters
Source: Archives of Environmental Health
Publication date: 2001-05-01
Arrival time: 2001-08-11


ABSTRACT. Several epidemics of nicotine intoxication have been described among tobacco harvesters; however, little is known about nicotine absorption under typical working conditions. To assess systemic nicotine absorption during a regular working shift, the authors performed an observational field study. Included in the study were 10 healthy, nonsmoking, female tobacco harvesters and a control group of 5 healthy, nonsmoking, female hospital workers. Nicotine and cotinine were measured in sequential samples of blood and urine during a regular workshift. Blood nicotine levels rose from a nadir value of 0.79 +/- 0.12 ng/ml to a peak value of 3.45 +/- 0.84 ng/ml (p < .05 [Tukey's modified t test]) in the exposed group. In the control group, levels were stable at 0.1 +/- 0.1 ng/ml (p < .01). Moreover, the mean blood nicotine level measured 3 mo following the end of exposure in 6 of 10 exposed subjects was 0.24 +/- 0.12 ng/ml (p < .01). Corresponding higher values of urine nicotine and urine cotinine were observed in the exposed versus control group (comparative p values were < .01 and < .05, respectively). Overall, tobacco harvesters absorbed approximately 0.8 mg of nicotine daily. Given that nicotine can induce adverse health effects, the authors believe that prevention of nicotine absorption in tobacco harvesters should be sought and that workers should be informed about occupational risks.

TOBACCO HARVESTERS may suffer an acute selflimited illness known as green tobacco sickness (GTS).1,2 The disease, characterized by vomiting, headache, abdominal pain and cramps, salivation, and sweating,3 occurs often among nonsmoking harvesters, particularly when they harvest wet tobacco.4 High nicotine levels5 in dew collected from tobacco leaves and high cotinine levels in urine of exposed workers have led investigators to hypothesize that GTS is a form of nicotine intoxication that results from percutaneous absorption of nicotine.6 Nicotine is an alkaloid that causes stimulation of autonomic ganglia and the central nervous system.7 Nicotine as a free base easily crosses the dermal barrier,8,9 and workers harvesting tobacco leaves can absorb it through the skin. Although more than 30,000 workers are employed every year in Italy'10 in tobacco farming, a direct measure of nicotine absorption during a regular workshift has never been reported.

To assess nicotine absorption among tobacco harvesters, we measured nicotine and cotinine concentrations in blood and urine samples collected before, during, and after a regular workshift. To estimate the external dose of exposure, we applied paper pads on the skin of each worker for later nicotine analysis.

Materials and Method

Study protocol. We studied 10 healthy, nonsmoking tobacco harvesters and 5 healthy, nonsmoking unexposed controls. We measured nicotine and cotinine levels in plasma and urine before, during, and after a regular workshift. To ascertain carry-over effects from previous days of work, we collected baseline urine and blood samples 3 mo after the end of the harvesting season from 6 of the 10 exposed subjects.

Study location. The study was performed in Umbria, central Italy, on October 1, 1997. This day occurred during mid-season for tobacco harvesting and in the middle of a work week. Umbria is a region with a high prevalence of tobacco farming. The farm that participated in the study is one of the largest in the region, employing approximately 400 workers each year. The farm tobacco practice is similar to that followed throughout Italy. The variety of tobacco harvested was Mcwrite.

Study population. Twelve healthy female nonsmokers who met all the inclusion criteria were selected through advertisements posted in the farm office. However, after the interview and the physical exam, 1 subject was excluded because she was pregnant and 1 could not participate because she was ill on the day of study.

The inclusion criteria were between 20 and 50 yr of age and general good health. Exclusion criteria were history of liver or skin disease, drug or alcohol abuse, and use of medication that might influence nicotine absorption (e.g., nitrates) or nicotine metabolism (e.g., barbiturates). All subjects answered a questionnaire about personal habits (previous smoking habits, exposure to environmental tobacco smoke at home), medical and occupational histories, and previous occurrence of symptoms of GTS. Five healthy, nonsmoking female volunteers were selected as nonexposed subjects among the hospital staff (research physicians).

Environmental sampling. On the day of the study, 50 ml of dew were collected from tobacco leaves and refrigerated for subsequent nicotine assay, Leaves from different sites of the field were collected, stored in plastic bags, and refrigerated for later nicotine assay.

Personal passive sampling. Paper pads (5 cm^sup 2^) were applied with tape directly on the skin of the torso of each worker. To avoid direct contamination with dew or tobacco leaves, we applied the pads under the clothes. Pads were applied before the beginning of the shift and removed at the end of the shift. Following removal, pads were placed individually in closed containers and refrigerated for subsequent nicotine analysis.

Biological sampling

Urine. Collection of urine was begun the night before the study and completed the night after the study at 12 P.M. Five urine specimens were collected from each subject in acidified plastic containers. We instructed exposed and nonexposed subjects to keep containers away from tobacco and tobacco smoke. Urine samples were brought to the lab and frozen at -70 deg C for subsequent analysis. Three months following the end of the harvesting season, we collected early morning urine samples from 6 of the subjects who had participated in the earlier study.

Blood. Blood was collected by phlebotomy from the antecubital vein. The first sample was collected before the shift; subsequent samples were collected about every 1 and 1/2 hr during the shift. The last sample was collected 2 hr after the shift had ended. We used the same time schedule to collect blood from the 5 nonexposed subjects. Following collection, blood samples were stored in ice, brought to the lab, and centrifuged, and serum was stored at -70 deg C for later analysis. Three months after the end of harvesting season, we collected one early morning blood sample from 6 of the subjects who participated in the study.

Exposure protocol. On the day of the study, subjects wore what they normally wore to work: rubberized apron, gloves and boots layered above cotton clothes. Subjects began working at 7 A.M. and worked until 10.00 A.M., at which time they paused for a 30-min breakfast break and shed protective rubberized clothes. They resumed work at 10:30 A.M. and continued until 2 P.M. Following a 2-hr lunch break, the subjects returned at 4.00 P.M. for the final blood sampling. Three months after the end of harvesting season, we collected early morning blood and morning urine samples from each of 6 of the 10 tobacco harvesters.

Control protocol. The same sampling protocol used with the tobacco harvesters was used for the 5 healthy volunteers who worked in our hospital. We asked the subjects to avoid exposure to environmental tobacco smoke during working hours.

Nicotine and cotinine assays. Nicotine and cotinine were determined in blood and urine by gas chromatography-mass spectroscopy. The limit of quantitation of the method is 0.20 ng/ml in plasma and urine.11 Nicotine from tobacco leaves and pads was extracted by the supercritical fluid method and then assayed by highperformance liquid chromatography.12,13

Statistical analysis. Statistical analysis was performed with SAS software (version 6.12). To evaluate the change in nicotine and cotinine measurements over time, we performed two-way repeated measures analysis of variance (ANOVA), with grouping by environmental tobacco smoke (ETS) exposure (present vs. absent). We then used Tukey's modified t test to perform pairwise comparisons. To compare tobacco harvesters with controls, we repeated the two-way repeated measures ANOVA with grouping by subject status (harvester vs. control).

Results

On the day of the study, the atmospheric conditions, which were typical for the area, were cold and foggy early in the morning. Tobacco leaves were covered with dew. Later during the day, it was sunny and warm, and no visible water remained on the leaves.

After the study, we found that 1 subject who smoked up until 1 wk before the study had very high plasma and urine cotinine values. Her cotinine data had to be excluded from the analysis. One other subject had problems with phlebotomy, and only 3 samples could be drawn from her.

Characteristics of study population. The average ages of the exposed and nonexposed group were 49 +/- 10 yr and 32 +/- 3 yr, respectively. All exposed and nonexposed subjects were lifetime nonsmokers, except 1 individual who had smoked up until 1 wk before the study and 1 of the nonexposed individuals who had smoked until 1 mo before the study. Six subjects in the exposed group and 3 in the nonexposed group were exposed to ETS at home where one or morefamily members were smokers. None of the subjects took medication that interfered with the absorption or metabolism of nicotine.

Environmental sampling and personal passive sampling. The concentration of nicotine in dew collected from tobacco leaves was 1.85 (mu)g/l. Tobacco leaves had an average nicotine concentration of 19.6 mg/gm of dry matter (range = 9.74-41.61 mg/gm). Nicotine was measurable in all the personal sampling pads; the average amount of nicotine in the pads was 5.2 (mu)g (range = 0.5-19.5 (mu)g).

Plasma nicotine. Plasma nicotine concentrations for exposed and nonexposed individuals are shown in Figure 1. In the exposed group, nicotine concentration increased from a preshift value of 1.85 +/- 0.46 ng/ml to an endshift value of 3.45 +/- 0.84 ng/ml (p < .01; Table 1). Nadir and peak values were observed at 10.30 A.wt. (i.e., before the breakfast pause) and at 4 P.M. (end of work day), respectively. Individual plots of the blood nicotine curves showed similar results. The difference between baseline and peak nicotine concentration for each subject is presented in Figure 2. No differences were observed between ETS-exposed and nonETS-exposed individuals, either at baseline or at any time point (Table 11.

Plasma nicotine values in the nonexposed group were about 6 times lower than plasma nicotine values in the exposed group (p < .01). No trend was observed during the sampling times. Baseline blood nicotine, measured in 6 of the exposed subjects 3 mo after the end of the harvesting season, was 0.24 +/- 0.12 ng/ml (n = 6); this level was significantly lower than the last value measured during the work day (Table 1).

Plasma cotinine. Plasma cotinine concentrations in exposed and nonexposed individuals are shown in Figure 3. No trend was observed across the shift during exposure (p = .22). No differences were observed between ETS-exposed and non-ETS-exposed subjects, either at baseline or at any time point (Table 1). The average plasma cotinine concentration measured in 6 of the exposed subjects 3 mo after the end of the harvesting season was 0.72 +/- 0.19 ng/ml (Table 1). The difference between exposed and control individuals did not reach statistical significance (p = .1).

Fig. 1.

Table 1.

Fig. 2.

Urine nicotine and cotinine levels. Urine nicotine and cotinine concentrations versus time for exposed and nonexposed individuals are shown in Figures 4 and

5. In the exposed group, there was a trend toward increasing urine nicotine (p = .098) and cotinine levels (p = .10) during the sampling period (Table 1), but no such trend was present in the nonexposed group. The difference between urine nicotine and cotinine levels in the exposed versus nonexposed group was statistically significant (p < .01 and p < .05, respectively). The average urine nicotine and cotinine concentrations measured in 6 of the exposed subjects 3 mo following the end of the harvesting season were 14.1 +/- 11.6 ng/ml and 6.19 +/- 1.13 ng/ml, respectively. These values were significantly lower than the last values measured during the work day (p < .05; Table 1).

Discussion

In this article, we have presented novel data on plasma levels of nicotine during tobacco harvesting. In other studies, investigators have reported increased urinary excretion of nicotine and cotinine following harvesting. These studies, however, did not provide information on the time course of exposure to nicotine during a work day. Our analysis also provided a quantitative estimate of nicotine exposure in tobacco harvesters.

We compared nicotine and cotinine levels in nonsmoking tobacco harvesters with nonsmoking women who did not work with tobacco. As an additional control, we compared levels of nicotine and cotinine in a group of tobacco harvesters while harvesting and again-3 mo after the harvest season ended. Both control groups had extremely low levels of nicotine and cotinine, compared with the tobacco exposure group, thus confirming that levels measured during harvesting were a consequence of occupational exposure.

Plasma nicotine levels in the exposed group rose significantly during the day and peaked at the end of the work day--a fact consistent with ongoing workplace exposure. Six of the study subjects were exposed at home to ETS. Given that the baseline value was the same in those with and without passive smoke exposure, one can assume that all subsequent increases in nicotine levels resulted from workplace exposure. We were, however, surprised to observe baseline nicotine levels among all the workers. Based on the short halflife of nicotine, one would not expect baseline levels to be explained by a carry-over effect from previous days of work. Nor were the workers exposed to ETS immediately before the shift commenced. Perhaps the rain gear (i.e., rubberized boots and apron) that each worker wore early in the morning was contaminated with residual nicotine from previous days. Indeed, baseline plasma nicotine measured at the end of harvesting season was approximately 10 times lower than the value measured the day of the study.

Fig. 3.

Fig. 4.

It is also interesting that the nadir value of plasma nicotine for the work group was reached at about 10:30 A.M., and it increased after workers shed their protective rubberized clothes. Although early in the morning the field was wet and leaves were moist with dew, there was little nicotine absorption, which occurred when workers shed protective clothes at a time when the leaves and field had been dried by several hours of sunshine. This result indicates that dermal absorption, rather than inhalation, is the main route for nicotine absorption in the workplace; that nicotine absorption occurs, even in the absence of visible moisture; and that rubberized clothes effectively protect workers from absorption. The efficacy of rubberized clothing and gloves in protecting workers from nicotine absorption has been shown in other studies.14,15 The measurement of nicotine on personal monitoring pads also supports this route of exposure. Nicotine inhalation cannot be excluded; however, if inhalation is an important route, one would expect to observe some nicotine absorption during the first hours of exposure, when dermal absorption is negligible, because workers are protected by rubberized clothing-but inhalation could still occur.

Fig. 5.

Plasma cotinine levels in exposed workers were about 50 times higher than in nonexposed workers. The values were rather stable during the workshift (i.e., averaging about 15 ng/ml). A flat cotinine curve reflected the long plasma half-life of cotinine (i.e., approximately 20 hr). Urine cotinine levels paralleled plasma cotinine levels. One can use steady-state plasma cotinine levels to estimate daily systemic intake of nicotine.16 Based on population data for the extent of conversion of nicotine to cotinine and for the clearance of cotinine, the following equation computes nicotine intake from steady-state cotinine levels: Dnic(mg) = Ccotcss(ng/ml) x 0.08, where Dnic is the daily dose of nicotine, and Ccotcss is the steady state plasma cotinine concentration.16

In our subjects, plasma cotinine averaged about 15 ng/ml, indicating a daily intake of 1.2 mg of nicotine. The comparison of the ETS-exposed group with the non-ETS-exposed group once again revealed no difference in plasma cotinine values--either at baseline or at any other time point--indicating that exposure at work was the major contributor to plasma cotinine in the exposed group.

Urine nicotine tended to increase throughout the work day, but this change was not significant, given the high variability observed.17,18 In addition, urine cotinine increased nonsignificantly across the workshift. Urinary excretion of cotinine was highly correlated with plasma cotinine value. Nicotine and cotinine levels measured in our study were approximately one order of magnitude lower than those observed in other studies of workers with GTS.19-21 Most of the previous studies, however, were performed in the United States and India during the late 1970s or early 1980s, and some of the differences might reflect an overall improvement of current working conditions. Another possible explanation is the different farming practices or personal behaviors that might occur among farmers from different countries.22,23 Finally, we studied professional tobacco harvesters who were less likely to be exposed to excessive amounts of nicotine than were inexperienced workers who were studied by other investigators.

We attempted to estimate the external dose of exposure by applying paper pads as a personal passive sampler. Paper pads have long been used as personal passive samplers to monitor pesticide exposure of farmers.24 Nicotine is absorbed through the skin; therefore, we decided to monitor nicotine exposure by applying pads directly on the skin below clothing. The amount of nicotine recovered from the pads was highly variable within the group and was not correlated with any other absorption indicator. It is possible that different behaviors of the study subjects might have occasionally contaminated the pads by direct contact with tobacco leaves.

In this study we describe, for the first time, a complete profile of nicotine and cotinine absorption and excretion in tobacco harvesters. However, there are some limitations that merit consideration. Urine containers for after-shift urine collection were given to subjects and kept in their houses until the following day. Although we advised them to avoid exposing containers to tobacco smoke, some contamination could have occurred. The diets of our subjects were not standardized. Nicotine is present in some vegetables and beverages25; therefore, some influence of the diets on the values observed cannot be excluded. Nonetheless, the low values in women who did not harvest tobacco and values in tobacco harvesters 3 mo following the end of the harvesting season provided evidence that the\re was no significant contribution of nicotine from food.

In conclusion, the results of our study indicated that harvesting tobacco in Italy exposed workers to a daily absorption of nicotine that is equivalent to smoking 1 cigarette. Their cumulative nicotine exposure was equivalent to 180 cigarettes per season and 5,000 cigarettes during their working life. We find it difficult to estimate the health effects of this nicotine absorption by comparison to smokers because, unlike smokers, tobacco workers are not exposed to tar, carbon monoxide, and other combustion products of smoked tobacco.26 Nicotine per se stimulates the sympathetic nervous system, increases heart rate and blood pressure slightly, and may adversely affect lipid metabolism and endothelial function. It is unclear whether these effects contribute significantly to tobacco-related cardiovascular disease. Perhaps of more concern for women who harvest tobacco is the possibility of nicotine toxicity during pregnancy.27 Nicotine exposure in rodents during pregnancy causes impaired fetal brain development28,29 and neonatal behavioral abnormalities.30 We do not know if the low levels of nicotine found in our tobacco workers were toxic to their cardiovascular and reproductive systems.

The authors thank Dr. Peyton Jacob for supervising the analytical chemistry analyses and Silvia Wu and Lisa Yu for performing analytical chemistry measurements.

This study was supported, in part, by the United States Public Health Service Grants DA02277 and DA01696 awarded by the National Institute on Drug Abuse.

Submitted for publication April 17, 2000; revised; accepted for publication January 15, 2001.

Requests for reprints should be sent to Alessandra d'Alessandro, M.D., Istituto di Medicina del Lavoro, Via E. dal Pozzo, Perugia 06122 Italy.

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24. Aprea C, Sciarra G, Sartorelli P, et al. Esposizione a fenitrothion durante la spillatura di piante ornamentali in serra. [Phenitrothion exposure during preparation of ornamental plants in greenhouse.] Folia Med 1996; 67(2):421-29.

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ALESSANDRA D'ALESSANDRO Dipartimento di Medicina Clinica e Sperimentale-Istituto di Medicina del Lavoro University di Perugia Perugia, Italy

NEAL L. BENOWITZ

Division of Clinical Pharmacology and Experimental Therapeutics University of California San Francisco, California

GIACOMO MUZI

Dipartimento di Medicina Clinica e Sperimentale-Istituto di Medicina del Lavoro University di Perugia Perugia, Italy

MARK D. EISNER

Division of Occupational and Environmental Medicine and Division of Pulmonary and Critical Care Medicine University of California San Francisco, California

SABRINA FILIBERTO

Dipartimento di Medicina Clinics e Sperimentale-Istituto di Medicina del Lavoro Universita di Perugia Perugia, Italy

PAOLO FANTOZZI LUIGI MONTANARI

Dipartimento di Scienze e Tecnologie Alimentari e della Nutrizione Istituto di Industrie Agrarie UniversitA di Perugia Perugia, Italy

GIUSEPPE ABBRITTI

Dipartimento di Medicina Clinica e Sperimentale-Istituto di Medicina del Lavoro Universita di Perugia Perugia, Italy

Copyright HELDREF PUBLICATIONS May/Jun 2001

Publication date: 2001-05-01
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