* Research Centre, Toxicology Division, Department of Internal Medicine, University of Pavia, Pavia, Italy, and
Research Centre, Toxicology Division, Salvatore Maugeri Foundation IRCCS, Institute of Pavia, Pavia, Italy
Received April 10, 2003; accepted May 27, 2003
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ABSTRACT |
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Key Words: ethanol; benzene; metabolism; bone marrow; phenol; catechol; hydroquinone; muconic acid.
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INTRODUCTION |
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Of concern are benzenes adverse effects on the hemopoietic system. Prolonged exposure results in severe blood dyscrasias in humans (Parke, 1996). Several studies have addressed the effects of benzene on hematopoietic stem cells and early progenitors; in rodents, benzene inhalation has been shown to decrease the number of early and/or late erythroid progenitors (BFU-E and CFU-E) and/or the myeloid progenitor cells (e.g., CFU-GM) (Baarson et al., 1984
; Corti and Snyder, 1996
; Dempster and Snyder, 1991
; Farris et al., 1997
). Although the mechanism of benzene toxicity is unclear, it is well established that benzene metabolism is required.
The ethanol-inducible cytochrome P450 2E1 (CYP2E1) accounts for the main fraction of hepatic and pulmonary benzene metabolism (Guengerich et al., 1991; Powley and Carlson, 2001
). Liver CYP2E1 catalyzes benzene oxidation to the reactive intermediate benzene oxide (Lindstrom et al., 1997
). Three main pathways have been identified downstream of the benzene oxide-oxepin. The first one leads to ring-hydroxylated compounds, such as phenol (Ph), catechol (Cat), hydroquinone (HQ), and 1,2,4-trihydroxybenzene, which are mostly excreted in the urine as sulphates and glucuronides (Sabourin et al., 1988
). These conjugates are regarded as detoxification products, because conjugation accelerates their elimination and prevents the formation of odd electron reactive molecules (Snyder et al., 1993
). The second pathway involves the ring opening and generates trans,trans-muconaldehyde and its corresponding final urinary metabolite trans,trans-muconic acid (MA) (Witz et al., 1989
). Additionally, variable amounts of different mercapturic acids, deriving from benzene oxide through conjugation routes involving glutathione (GSH), are excreted in human and animal urine (Snyder et al., 1993
).
Reactive benzene intermediates are transported via the blood from the liver and lungs to the bone marrow, where they accumulate (Rappaport et al., 2002). Here, further transformations of benzene oxide and metabolites can occur, including the conversion of dihydroxy metabolites (HQ, Cat, 1,4-, and 1,2-dihydrodiols and diolepoxides) to the corresponding quinones and semiquinones (Golding and Watson, 1999
; Oshiro et al., 2001
). All such molecules and trans,trans-muconaldehyde have been shown to form adducts to nucleic acids and proteins in vitro and possibly cause cell damage in vivo (Snyder, 2002
). Benzene myelotoxicity has been suggested to result from the interactive effects of several metabolites that produce alterations by radical-mediated reactions or by supporting radicalic processes of reactive oxygen species (Snyder, 2002
; Witz et al., 1996
).
The modulation of benzene metabolism can modify benzenes excretion profile and/or toxicity. For example, CYP2E1 knock-out mice displayed lower excretions of benzene urinary metabolites, cytotoxicity, and genotoxicity than did wild-type mice (Valentine et al., 1996), and microsomal epoxide hydrolase-deficient CD-1 mice were almost unresponsive to benzene-induced genotoxicity (Bauer et al., 2003
). On the other hand, in rats exposed to 500 ppm of benzene (2 h/day for 3 weeks), repeated ingestion of ethanol, a potent inducer of CYP2E1-mediated benzene metabolism (Sato et al., 1980
), accelerated benzene disappearance from the blood, decreased urinary Ph excretion, and aggravated benzene-induced hemopoietic disorders (Nakajima et al., 1985
). In other studies in rodents, ethanol worsened the benzene-induced reduction in bone marrow and spleen cell number and increased the severity of anemia and lymphocytopenia (Baarson et al., 1982
). Although several experiments have documented the role of ethanol in benzene toxicity (Baarson and Snyder, 1991
; Baarson et al., 1982
; Nakajima et al., 1985
; Seidel et al., 1990
), these combined exposure studies had, with few exceptions (Corti and Snyder, 1996
; Daiker et al., 2000
), the limitation of using benzene doses that were much higher (
300 ppm) than those relevant to human exposures. In this respect, while much is known about the effects of benzene poisoning, the mechanisms by which lifestyle and environment factors may influence the individual response to low-level benzene exposure are still poorly understood.
This study was designed to assess the hypothesis that cotreatment with ethanol would affect benzene myelotoxicity and urinary biomarkers of benzene exposure. In particular, the purpose of the experiments was 3-fold: to assess (1) the effect of repeated alcohol ingestion on the myelotoxicity of 10 ppm benzene in mice; (2) the relation existing between benzene + ethanol-induced myelotoxicity and the urinary levels of benzene metabolites, namely, Ph, Cat, HQ, and MA; and (3) if measurements of urinary metabolites that are commonly used as indicators of benzene exposure are reliable when applied to animals coexposed to low levels of benzene and ethanol.
In the past, the analysis of urinary phenolic compounds has been widely used to assess occupational exposure to benzene at levels above 5 ppm (Qu et al., 2000). More recently, urinary MA has been introduced to monitor exposure to lower benzene concentrations (Boogard and van Sittert, 1995
). Studies in this field indicated that several endogenous, environmental, and dietary factors can influence benzenes urinary metabolic profile (Marrubini et al., 2002
; McDonald et al., 2001
).
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MATERIALS AND METHODS |
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Analytical-grade N,N-dimethylformamide (99% purity), pure trifluoroacetic acid (TFA, 99% purity), concentrated hydrochloric acid (densimetric grade 1.18 g/ml or about 37%, w/w), and sodium hydroxide pearls were purchased from BDH (Milan, Italy).
A concentrated enzyme solution of ß-glucuronidase-arylsulphatase type H-1 from Helix Pomatia (cat. n. 127060) was obtained from Roche Diagnostics (Milan, Italy). Trans,trans-muconic acid (98% purity), catechol (99% purity), hydroquinone (99% purity), and phenol (>99.5% purity) were provided by Sigma-Aldrich (Milan, Italy).
HQ, Cat, and Ph stock solutions (20 mg/ml each) were prepared in methanol, while that of MA (0.5 mg/ml) was prepared in N,N-dimethylformamide-methanol (1:10, v/v). All of the solutions were stored in the dark at -20°C. For spiking or HPLC analysis, each stock solution was diluted with 0.01-N aqueous HCl just before use.
Animals
Male CD-1 mice weighing 25 to 30 g were purchased from Charles River Italia (Calco, Lecco, Italy). They were housed under controlled conditions of temperature and humidity. They were acclimated for 1 week to a 12-h light12-h dark cycle and maintained on food and water ad libitum prior to their inclusion in the experiments. Thereafter, the mice were randomly assigned to four experimental groups (n = 8): control animals, those treated with ethanol, those treated with benzene, and those treated with both. All of the experiments were conducted in compliance with the European Community Guide for the Care and Use of Laboratory Animals (European Council Directive 24-11-1986, 86-609-EEC).
Treatments
Ethanol.
Mice of the ethanol and the ethanol + benzene groups were fed the Lieber-DeCarli liquid diet containing 5% (w/v) ethanol (corresponding to 36% of energy as ethanol) for 3 weeks (weeks 24). Ethanol was gradually introduced in the diet to reach the 5% concentration over a period of 7 days on week 1, starting with 3% (w/v) on the first 2 days, 4% for the next 2 days, and the final concentration of 5% thereafter. Concomitantly, the control and the benzene groups were administered the Lieber-DeCarli isocalorically balanced dextrin-maltose diet for 4 weeks. The daily average caloric and alcohol intakes were 400500 kcal/kg body weight (bw) and 2025 g ethanol/kg bw, respectively. Ethanol was administered in a Lieber-DeCarli liquid diet in order to achieve enzymatic induction of CYP2E1 without causing fasting or a lower daily intake of carbohydrate (Daiker et al., 1999; Lieber et al., 1989
). These conditions are recognized factors other than ethanol that induce CYP2E1 (Johansson et al., 1988
), but they are less controllable.
Benzene.
In order to mimic the heavy daily human intake of benzene on working days, the animals were exposed to 10 ppm benzene vapors, 6 h daily, 5 days per week, for 2 weeks (weeks 34). In workplaces, 1 ppm is the current permissible exposure limit (PEL), with a short-term exposure limit (STEL) of 5 ppm averaged over 5 min. However, these limits are often exceeded in developing countries (Rothman et al., 1998).
In the present study, benzene exposure was carried out in a dynamic exposure chamber of 1 m3. The air supply was passed through activated carbon and high-efficiency particulate air (HEPA) filters. The benzene concentration in the chamber was continuously monitored by an automatic benzene, toluene, and xylene portable gas chromatograph equipped with a flame ionization detector BTX61M. The groups of mice, fed either the ethanol or dextrin-maltose diet and exposed to air in an identical exposure chamber, served as the controls.
All of the mice were sacrificed by cervical dislocation immediately after the last exposure to benzene or air.
Myelotoxicity Assessment
Isolation of murine bone marrow cells.
Immediately after sacrifice, the intact femurs were isolated, cleaned, and cut at both ends just below the head. Each femurs marrow was flushed with 3-ml IMDM under sterile conditions. A single-cell suspension was produced by repeatedly drawing the cells through a syringe fitted with a 23-gauge needle, followed by filtration through a 100-µm cell strainer. After washing by centrifugation, the cells were resuspended in IMDM supplemented with 30% FBS, counted in a hemocytometer, and diluted properly to be used in the colony-forming assays.
Bone marrow colony-forming units: Erythroid and burst-forming unit-erythroids.
Two erythroid progenitor cell types have been identified, that is, the more differentiated colony-forming unit-erythroid (CFU-E) and the less differentiated burst-forming unit-erythroid (BFU-E). These cell types can be distinguished on the basis of their microscopic appearances and by varying the duration of the culture. The growth of CFU-E and BFU-E was supported in vitro by means of a methylcellulose semisolid culture medium (Methocult M3334) containing 3 units/ml rh erythropoietin as described in Malerba et al. (2002). A 300-µl sample of an individual mouse bone marrow cell suspension was added to a 4-ml aliquot of culture medium at the final concentration of 300,000 cells/ml. The cellmedium mixture was dispensed in 1-ml aliquots in three 35-mm petri dishes. The cultures were incubated at 37°C under saturated humidity. The erythroid colonies were scored under an inverted microscope after either 3 days (CFU-E) or 9 days in culture (BFU-E), respectively.
Bone marrow colony-forming units: Granulocyte macrophage.
Granulocyte macrophage colony-forming units (CFU-GM) were assayed as described in Malerba et al. (2002), using the methylcellulose semisolid medium Methocult GF M3534. The medium contained 10-ng/ml IL-3, 10-ng/ml IL-6, and 50-ng/ml SCF as a source of colony-stimulating factors. Each 35-mm culture dish contained 50,000 bone marrow cells/ml methylcellulose medium, and each sample was cultured in triplicate. The cultures were incubated for 7 days at 37°C. On microscopic examination, aggregates containing more than 50 cells were defined as colonies.
Urinary Benzene Metabolites
The urine samples were collected in metabolic cages during the last 6-h benzene or air-exposure session, at the end of each week (weeks 14).
Benzene metabolites were measured directly in the urine by using a coupled-column reversed-phase liquid chromatographic analyzer with ultraviolet (UV) and fluorimetric (FL) detection (LC/LC-UV-FL), as described in Marrubini et al. (2001).
Sample treatment for MA determination.
Aliquots of 0.10.25 ml of urine samples were diluted (1:1, v/v) with 0.05-N aqueous HCl, vortexed, and centrifuged at 4500 rpm for 15 min. One hundred µl of the resulting clear solution at pHs between 1 and 2 were injected into the LC-LC analyzer.
Sample treatment for HQ, Cat, and Ph determination.
Aliquots of 0.1 ml of urine were mixed with 0.38 ml of 0.2-M sodium acetate buffer (pH 5) and 0.02 ml of ß-glucuronidase-arylsulphatase concentrated solution and put in a thermostatic bath at 37°C for 18 h. On the following day, the sample was divided into four 0.1-ml aliquots that were immediately stored in the dark and frozen at -80° C. One aliquot per each analyte was thawed just before analysis, homogenized by vortex-mixing for a few seconds, and centrifuged at 4500 rpm for 15 min at 14°C. Twenty-five µl of the supernatant were directly injected into the analyzer.
Sample analysis.
The LC-LC-UV procedure adopted for MA analysis has been described in detail in Marrubini et al. (2001). For HQ, Cat, and Ph direct analysis, modifications were introduced in the cited procedure and three single-residue LC-LC-UV-FL methods were developed (unpublished data).
Data analysis.
All of the data are expressed as mean ± SD. Statistical significance was assessed by one-way ANOVA followed by Fishers PLSD for comparison of more than two groups (according to the different types of treatments). To determine the differences within each treated group, a statistical analysis of the results was performed by means of repeated measures ANOVA. A value of p < 0.05 was considered statistically significant.
Bone marrow colony-forming units of raw data were expressed as number of colonies/dish. The mean colony count of three dishes was calculated for each mouse, and the mean value per each treatment group was determined. The mean value obtained in the control group was taken as 100%, and the values obtained after ethanol, benzene and ethanol + benzene treatments were expressed as % ± SD of the control group.
The metabolites urinary concentrations measured were expressed as µg/l urine on the basis of the amount of urine (in l) excreted by each animal during the exposure session. The data, expressed as µg/kg bw, were calculated considering the bw (in kg) of each mouse.
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RESULTS |
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Urinary Benzene Metabolites
Considerable "background" concentrations of Ph and other benzene metabolites were found in urine of the control, untreated animals. Similar findings have been previously reported in rats (Marrubini et al., 2002) and humans (Qu et al., 2000
). The origin of these benzene-derived compounds in "nonexposed" individuals is still unclear.
Phenol.
In mice inhaling benzene a significant increase in Ph was observed: Urinary Ph levels, ranging between 2200 to 2400 µg/kg bw before benzene exposure, reached 4931 ± 1055 at week 3 and then declined to 3909 ± 984 µg/kg at week 4 (Fig. 2). The mice that were given ethanol alone showed a slightly lower, though not statistically significant, excretion of Ph at all time points, compared with the controls.
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Catechol.
In the mice treated with 10 ppm of benzene, the urinary excretion of Cat was significantly increased as compared to the background values measured in the preexposure period (Fig. 3).
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Hydroquinone.
Urinary HQ levels (538 µg/kg bw) significantly exceeded background values in mice exposed to benzene alone. The HQ elevation was much more pronounced at the end of the first week (week 3: 739 ± 137 µg/kg bw) than after the second week (week 4: 337 ± 72 µg/kg bw) of benzene treatment (Fig. 4).
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Trans,trans-muconic acid.
In mice treated with benzene only, the urinary concentrations of MA measured at weeks 3 and 4 were 25- and 10-fold higher, respectively, than in the preexposure period (Fig. 5). Again, the elevation of the metabolite levels was more pronounced at week 3 compared with week 4. Ethanol treatment was per se devoid of any effect on the urinary levels of MA (Fig. 5
, compare "Control" line with "Ethanol" line) but almost completely suppressed the increase in urinary MA excretion in the benzene-exposed mice (Fig. 5
, "Ethanol + Benzene").
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DISCUSSION |
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Mice inhaling benzene alone (10 ppm, 6 h/day, for 2 weeks) showed no weight loss as compared with controls, while both groups of ethanol-fed animals significantly lost weight (Table 1). Since the overall food consumption decreases in rodents administered ethanol, this factor was controlled by pair feeding each control animal with amounts of the liquid diet equal to those ingested by the corresponding alcohol-treated littermates.
Our data showed a significant reduction in the CFU-E, BFU-E, and CFU-GM counts in mice inhaling 10 ppm of benzene, as compared with the untreated controls. Similar decreases in the in vitro colony-forming ability of CFU-E have been reported previously in DBA/2J male mice exposed to the same 6-h daily dose of benzene for 5 days (Dempster and Snyder, 1991) as well as in C57B1/6J male mice exposed for 32, 66, and 178 days (Baarson et al., 1984
). Baarson and coworkers also observed a depression of BFU-E to 55% of the control growth values after 66 days of 10 ppm benzene treatment. Consistent with other investigations (Dempster and Snyder, 1991
), we found a preferential susceptibility of the erythroid progenitor cells to benzene toxicity as compared to their granulocytic counterparts. On the other hand, no reduction in cell numbers was reported by Farris and coworkers (1997)
in experiments using the CFU-E and CFU-GM assays in B6C3F1 mice given 10 ppm benzene (6 h/day, 5 days/week, up to 8 weeks of exposure). This difference may indicate a lower susceptibility of B6C3F1 mice to benzene toxicity compared with the CD-1 strain, as is also suggested by studies showing a similar difference of these two strains in response to the genotoxic effects of trans,trans-muconaldehyde (Oshiro et al., 2001
).
In the present study, ethanol consumption enhanced benzene-induced bone marrow cell damage, in terms of depression of the number of erythroid and granulocyte-macrophage progenitors. Notably, ethanol itself impaired the ability of CFU-E, BFU-E, and CFU-GM to form colonies in vitro, and the ethanol-benzene combination resulted in more severe myelotoxicity than benzene itself (Fig. 1).
In adult C57B1/6J male mice, the concomitant ingestion of ethanol (5 or 15% in drinking water 4 days/week) with 300 ppm benzene (6 h/day, 5 days/week, 13 weeks) was shown to exacerbate benzene-induced anaemia, lymphocytopenia, and depression of marrow and splenic cellularities (Baarson et al., 1982). In the same animal model, coexposure to 5% ethanol and 300 ppm benzene (6 h/day, 5 days/week, 9 weeks) reduced the number of CFU-E to a greater extent than exposure to benzene alone. The BFU-E cell numbers were similarly decreased in benzene and ethanol-benzene treated mice, both being around 12% of the controls (Baarson and Snyder, 1991
).
At much lower exposure levels (10 ppm benzene, 6 h/day over 10 consecutive days), Swiss Webster adult male mice presented severe CFU-E count depression (64% of the controls), but this effect was not worsened by the coadministration of ethanol (5% via drinking water) (Corti and Snyder, 1996).
Our study also examined the effect of benzene and ethanol on urinary levels of benzene metabolites measured at various time intervals during exposure. In mice inhaling benzene only, Ph was by far the most abundant urinary compound, followed by HQ, MA, and Cat. The concentration of hydrosoluble metabolites apparently peaked in the first week of exposure and decreased thereafter (Figs. 25
). The mechanism underlying the time-related change in urinary metabolite levels was not investigated. Initial tissue accumulation of reactive intermediates and their covalent attack on cellular components involved in benzene metabolism may have reduced enzyme activity. A process of this type was described by Costa et al. (1999)
in streptozocin-induced diabetic rats treated with benzene.
The destruction of CYP hemeprotein due to benzene and its major metabolites has been demonstrated in vitro (Soucek et al., 1994). In addition, benzene and the metabolites benzene oxide-oxepin, Ph, Cat, and HQ were shown to compete in vitro for the same CYP450 2E1 isozyme (Rothman et al., 1998
). Both of these processes may result in reduced levels of the metabolites requiring a second oxidation, that is, HQ and Cat (from Ph) and MA (from benzene oxide-oxepin). Urinary Ph levels can be expected to decrease to a lower extent, because benzene oxide may rearrange nonenzimatically to form Ph or is converted to Ph by enzyme activities not requiring CYP2E1mediated oxidation (Powley and Carlson, 2001
). Accordingly, from week 3 to week 4 of benzene exposure, the Ph levels decreased by little more than 25% (from 4931 ± 1055 to 3909 ± 984 µg/kg bw; Fig. 2
), while the levels of HQ and MA had around 50% decrement (from 739 ± 137 to 337 ± 72 µg/kg bw, and from 534 ± 92 to 243 ± 55 µg/kg bw, respectively; Figs. 4
and 5
). Cat was always excreted at low amounts, and no significant changes were observed in urinary levels of this metabolite from week 3 to week 4 (Fig. 3
). In humans, urinary Cat levels should be distinguishable from background values only at benzene doses higher than 510 ppm causing saturation of all of the faster metabolic pathways (Rothman et al., 1998
).
Ethanol administration was associated with significant changes in the urinary excretion of benzene metabolites. The type of response was somewhat unexpected. Ethanol remarkably reduced the urinary levels of all hydrosoluble metabolites (Ph, HQ, Cat, and MA) in mice showing enhanced benzene myelotoxicity.
As a possible explanation of this finding, one may suggest that chronic treatment with ethanol caused CYP2E1 induction, thus enhancing benzene oxidation to reactive derivatives, as supported by the hemotoxicity data. Because of extensive production of electrophilic intermediates, more macromolecular adducts and reactive oxygen species originated locally in the bone marrow, and this in turn caused damage to the bone marrow precursor cells. Concurrently, the predominant formation of reactive intermediates and their binding to cellular macromolecules may have reduced the amounts of free and conjugated metabolites available for urinary excretion. Several observations suggest that the extent to which any hydroxylated metabolite can be generated during benzene metabolism is related to the specific balance of reductases (NQO1, microsomal epoxide hydrolases, etc.) and oxidative enzymes (CYP2E1, myeloperoxidases, etc.) (Snyder, 2002). Ethanol could shift this balance toward a higher net amount of reactive metabolites, causing an increased production of macromolecular adducts in tissues and more severe cell damage in bone marrow. Recently, in workers exposed to benzene, a positive association was demonstrated between alcohol consumption and increased generation of albumin adducts to either 1,4-benzoquinone or benzene oxide as a consequence of CYP2E1 induction by ethanol (Rappaport et al., 2002
). However, no information is available on concentrations of phenolic metabolites in alcohol-consuming subjects.
An additional mechanism to explain the lower urinary concentration of metabolites in the coexposed animals could be a direct competition between ethanol, benzene, and its metabolites for CYP2E1. The inhibition of enzyme activity may occur following acute exposure to benzene and ethanol (Johansson et al., 1988; Mikov et al., 2000
; Sato et al., 1981
). In most cases, however, this interaction was observed in single-dose experiments using very high concentrations of benzene and ethanol. In principle, given the putative role of benzene metabolites as mediators of toxicity, the inhibition of benzene metabolism should result in reduced rather than increased myelotoxic damage. In fact, our experiments demonstrated a higher hemopoietic stress caused by the combination of ethanol and benzene compared to benzene alone. Studies have indicated that competitive inhibition for CYP2E1 (Tuo et al., 1996
), lack of CYP2E1 expression (Valentine et al., 1996
), and microsomal epoxide hydrolases deficiency (Bauer et al., 2003
) are protective factors from benzene-induced toxicity. If competitive inhibition took place under our experimental conditions, this process should have decreased the formation of myelotoxic metabolites (mainly benzene oxide, 1,4-benzoquinone, and trans,trans-muconaldehyde). Thus, other mechanisms are apparently involved in ethanol-induced potentiation of bone marrow damage in benzene-exposed animals.
Although the ethanol-supplemented liquid diet has been shown to prevent defective intake of nutrients (Lieber et al., 1989), in our experiments ethanol administration altered the bw gain, in accordance with the results of other studies in mice given comparable ethanol levels (Daiker et al., 1999
). Notably, ethanol-induced decreases in the urinary excretion of benzene metabolites, especially Ph and Cat, occurred not only in mice of the benzene group but also in those not receiving benzene. Ethanol consumption can interfere with nutrients, despite an adequate diet (Lieber, 2000
). Studies are needed to establish whether ethanol can affect the metabolism of dietary components or physiological precursors that, like benzene, are excreted in urine as MA or phenolic derivatives.
It may also be postulated that, in mice treated with ethanol, the metabolism of benzene was switched to alternative pathways. One pair of benzene metabolites that was not measured in our study, prephenyl- and phenylmercapturic acids, originates from the reaction of glutathione (GSH) with benzene oxide. In preliminary experiments, we found much lower urinary levels of both phenylmercapturic acid and MA in male Sprague-Dawley rats exposed to 1 ppm of benzene plus ethanol (given by a Lieber-DeCarli liquid diet) than in animals treated with benzene alone. It seems unlikely from these data that the metabolism of benzene was diverted to the GSH-dependent pathway due to the coadministration of ethanol.
A further hypothesis is that the action of ethanol was unrelated to alterations in benzene metabolism. Theoretically, ethanol could increase the exhalation of unmetabolized benzene and reduce benzene retention in the body by altering ventilation, organ blood perfusion, and cardiac output. However, Driscoll and Snyder (1984) reported no significant changes in pulmonary ventilation in mice coexposed to ethanol (10%) and benzene (300 ppm, 6 h/day for 20 days) relative to mice exposed to benzene alone.
In conclusion, our data show that repeated ethanol ingestion can exacerbate myelotoxicity and affect biological markers of benzene exposure, as indicated by a lower urinary recovery of benzene metabolites (MA and phenolic compounds) deriving from bioactivation pathways. It is well accepted that biomonitoring studies allow more valuable information on an individuals exposure to solvents than air concentration measurements, in that the former tests reflect metabolic and toxicokinetic processes. Our data suggest that alcohol intake may be a confounding factor when interpreting biological monitoring data of benzene exposure. In view of the wide consumption of alcohol, this factor needs to be considered carefully in studies of workers occupationally exposed to benzene.
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ACKNOWLEDGMENTS |
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This study was supported by a grant from the Italian Ministry of Health, which has no control over the resulting publication.
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NOTES |
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