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Acrylamide is a probable human carcinogen with widespread exposure via food. The present study compared acrylamide intake measurements obtained from haemoglobin adduct levels and self-registered dietary consumption data in a group of 144 Norwegian healthy adults. Acrylamide adducts to N-terminal valine in haemoglobin were measured and used to estimate the intake via the internal dose approach which showed a median (interquartile range) of 0.24 (0.19–0.30) μg/kg bw/day. Data from weighed food records and food frequency questionnaires from the same individuals were used for probabilistic modelling of the intake of acrylamide. The median acrylamide intake was calculated to be 0.26 (0.16–0.39) and 0.30 (0.23–0.39) μg/kg bw/day, respectively from the two sources of self-registered dietary consumption data. Overall, a relatively good agreement was observed between the methods in pairwise comparison in Bland-Altman plots, with the methods disagreeing with 7% or less of the values. The intake estimates obtained with the two dietary consumption methods and one biomarker method are in line with earlier dietary estimates in the Norwegian population. The Margin of Exposure indicate a possible health risk concern from dietary acrylamide. This is the first study with a comparison in the same individuals of acrylamide intake estimates obtained with these methods. [ABSTRACT FROM AUTHOR]

Acrylamide (ACR) is a 2 alkene monomeric reagentused to synthesize polymers for industrial application. Evaluating its haematological toxicity action has been a preponderant objective of worldwide concern since the chemical was detected in human umbilical cord blood. In this work, its toxicity (ACR) in adult zebrafish (Danio rerio) peripheral blood was tested maintaining the fishes during 8 days in a 0.50 mM ACR LC50 and normal controls. Haemoglobin (Hb) analysis was performed by cellulose acetate electrophoresis. The activity ofä-aminolevulinic acid dehydratase (ALAD) determination by spectrophotometric method was used as a toxicity biomarker. DNA analysis of nucleated erythrocytes in blood extensions contrasted with Feulgen staining was also performed. All these data were processed through the Fiji-ImageJ program platform and the GraphPad Prims statistic-7. The controls, from the peripheral blood electrophoretic profiles showed two main bands of globin, and one secondary minor band. Otherwise, those treated with ACR showed one main globin band and three minor bands, in the electrophoresis. The ALAD enzyme activity showed a 20% reduction compared with the controls. The erythrocytes presented nuclear DNA fragmentation in 80% of the analysed erythrocyte population, with respect to the control erythrocyte population. The haematocrit evaluation was higher in peripheral blood treated with ACR. The current study showed that ACR altered the structure of peripheral blood Hb, probably through the generation of ACR-Hb and / or ACR-GB type adducts, which may be expressed in the weakest bands found. In addition, it was possible to demonstrate that ACR can inhibit the enzymatic activity of ALAD, serving as a possible biomarker of haemotoxicity. The preceding was also accompanied by an apoptosis process evidenced through Feulgen staining. In conclusion, the ACR has haemotoxic activity in addition to the well-known neurotoxicity already reported, which could jeopardize the activity of oxygen supply in the blood and compromise processes in human foetal development. [ABSTRACT FROM AUTHOR]

Purpose: The main aim of this study was to investigate the cobalt (Co) concentrations in urine along 4 months and their relationship with Co concentrations in blood and haemoglobin (adducts) in 34 workers from a hard metal manufacturing plant where metallic Co and Co oxide were used. Furthermore, the excretion kinetics of Co was investigated and the half-lives of Co in blood, plasma and urine were calculated along 18 days of non-exposure in the same workers. Methods: Co was analysed, in all biological samples, by ICP/MS. Results: Wide fluctuations in the urinary Co concentration were observed throughout the work shift and during the work week. A highly significant linear correlation was found between Co concentration (geometrical mean) in urine samples provided each Thursday (end shift) during 16 subsequent weeks and levels of Co-haemoglobin adducts or blood Co concentrations at the end of the same period. The Co elimination kinetics in globin calculated along 18 days without Co exposure was slow, being related to the physiological metabolism of haemoglobin, while in blood, plasma and urine Co half-lives were 12.3, 9.1 and 5.3 days, respectively. Conclusion: Co concentrations in haemoglobin or blood are highly related to the geometrical mean concentration of urinary Co when samples are collected weekly for several subsequent weeks. The biological monitoring of occupational exposure to Co in hard metal facilities provides reliable results by using the Co concentrations in haemoglobin or in whole blood. The urinary findings, though, do not show the same reliability because of their wide daily and weekly fluctuations. [ABSTRACT FROM AUTHOR]

Background: Acrylamide is a chemical compound present in tobacco smoke and food, classified as a probable human carcinogen and a known human neurotoxin. Acrylamide is formed in foods, typically carbohydrate-rich and protein-poor plant foods, during high-temperature cooking or other thermal processing. The objectives of this study were to compare dietary estimates of acrylamide from questionnaires (DQ) and 24-h recalls (R) with levels of acrylamide adduct (AA) in haemoglobin. Methods: In the European Prospective Investigation into Cancer and Nutrition (EPIC) study, acrylamide exposure was assessed in 510 participants from 9 European countries, randomly selected and stratified by age, sex, with equal numbers of never and current smokers. After adjusting for country, alcohol intake, smoking status, number of cigarettes and energy intake, correlation coefficients between various acrylamide measurements were computed, both at the individual and at the aggregate (centre) level. Results: Individual level correlation coefficient between DQ and R measurements ( r) was 0.17, while r and r were 0.08 and 0.06, respectively. In never smokers, r, r and r were 0.19, 0.09 and 0.02, respectively. The correlation coefficients between means of DQ, R and AA measurements at the centre level were larger ( r > 0.4). Conclusions: These findings suggest that estimates of total acrylamide intake based on self-reported diet correlate weakly with biomarker AA Hb levels. Possible explanations are the lack of AA levels to capture dietary acrylamide due to individual differences in the absorption and metabolism of acrylamide, and/or measurement errors in acrylamide from self-reported dietary assessments, thus limiting the possibility to validate acrylamide DQ measurements. [ABSTRACT FROM AUTHOR]

N-Phenyl-2-naphthylamine (P2NA) was widely used as oxidation inhibitor, particularly in rubber manufacturing. Technical-grade P2NA was contaminated with carcinogenic 2-naphthylamine (2NA), and bladder cancer risk in exposed workers was attributed to this impurity. Investigations in humans and mammalian species revealed that small amounts of 2NA are excreted into urine after exposure to P2NA. However, since 2NA per se is not carcinogenic and main downstream metabolites of 2NA have not been found in urine so far, it remained uncertain if 2NA derived from P2NA dephenylation is further activated to carcinogenic downstream metabolites. An experimental animal study was therefore designed to indicate if, and if yes to which extent, 2NA from P2NA dephenylation is accessible to the metabolic pathway that is held responsible for the carcinogenicity of 2NA. Groups of 5 male and female CD rats were dosed with P2NA (2-550 mg/kg b.w.) and 2NA (0.075-75 mg/kg b.w.); 2NA-haemoglobin adducts and urinary 2NA excretion were determined applying GC-MS/MS. 2NA haemoglobin adducts originated dose-dependently after 2NA and P2NA dosing. To induce identical adduct concentrations, an approximately 100-200-fold higher dose of P2NA was necessary compared to 2NA. Since haemoglobin adducts are formed by the same pathway ( N-hydroxylation) as the ultimate carcinogens from 2NA, the comparison of adduct concentrations after 2NA and P2NA dosage permits a quantitative estimate of the carcinogenicity of P2NA. The results show that 2NA derived from dephenylation of P2NA enters the carcinogenic downstream pathway of 2NA in rats. Hence, the bladder cancer risk after human exposures to P2NA must be re-evaluated. [ABSTRACT FROM AUTHOR]

Abstract: Haemoglobin adducts are highly valuable biomarkers of cumulative exposure to carcinogenic substances. We have developed and applied an analytical method for the simultaneous quantification of five haemoglobin adducts of important occupational and environmental carcinogens. The N-terminal adducts were determined with gas chromatography as pentafluorophenylthiohydantoine derivatives according to the modified Edman-procedure and subsequent acetonization of the glycidamide adduct N-(R,S)-2-hydroxy-2-carbamoylethylvaline (GAVal). The use of self-synthesized labelled internal standards in combination with tandem mass spectrometry using negative chemical ionisation guarantees both high accuracy and sensitivity of our determination. The limit of detection for N-2-hydroxyethylvaline (HEVal), N-(R,S)-2-hydroxypropylvaline (HPVal), N-2-carbamoylethylvaline (AAVal) and N-(R,S)-2-hydroxy-2-carbamoylethylvaline (GAVal) was 2pmol/g globin, for N-2-cyanoethylvaline (CEVal) it was determined as 0.5pmol/g globin, which was sufficient to determine the background levels of these adducts in the non-smoking general population. The between-day-precision for all analytes using a human blood sample as quality control material ranged from 4.7 to 12.3%. We investigated blood samples of a small group (n =104) of non-smoking persons of the general population for the background levels of these haemoglobin adducts. The median values for HEVal, HPVal, CEVal, AAVal and GAVal in a group of 92 non-smoking persons were 18.1, 4.1, <0.5, 29.9 and 35.2pmol/g globin, respectively. The adduct levels in 12 persons reporting exposure to passive smoke at home were similar for most adducts with median values of 17.2, 4.1, 1.0, 24.9 and 29.7pmol/g globin for HEVal, HPVal, CEVal, AAVal and GAVal, respectively. Our results point to an elevated uptake of acrylonitrile caused by passive smoking as indicated by higher levels of the corresponding haemoglobin adduct CEVal. [ABSTRACT FROM AUTHOR]

In contrast to some extensively examined food mutagens, for example, aflatoxins, N-nitrosamines and heterocyclic amines, some other food contaminants, in particular polycyclic aromatic hydrocarbons (PAH) and other aromatic compounds, have received less attention. Therefore, exploring the relationships between dietary habits and the levels of biomarkers related to exposure to aromatic compounds is highly relevant. We have investigated in the European Prospective Investigation into Cancer and Nutrition (EPIC) cohort the association between dietary items (food groups and nutrients) and aromatic DNA adducts and 4-aminobiphenyl-Hb adducts. Both types of adducts are biomarkers of carcinogen exposure and possibly of cancer risk, and were measured, respectively, in leucocytes and erythrocytes of 1086 (DNA adducts) and 190 (Hb adducts) non-smokers. An inverse, statistically significant, association has been found between DNA adduct levels and dietary fibre intake (P = 0·02), vitamin E (P = 0·04) and alcohol (P = 0·03) but not with other nutrients or food groups. Also, an inverse association between fibre and fruit intake, and BMI and 4-aminobiphenyl-Hb adducts (P = 0·03, 0·04, and 0·03 respectively) was observed. After multivariate regression analysis these inverse correlations remained statistically significant, except for the correlation adducts v. fruit intake. The present study suggests that fibre intake in the usual range can modify the level of DNA or Hb aromatic adducts, but such role seems to be quantitatively modest. Fibres could reduce the formation of DNA adducts in different manners, by diluting potential food mutagens and carcinogens in the gastrointestinal tract, by speeding their transit through the colon and by binding carcinogenic substances. [ABSTRACT FROM PUBLISHER]

Abstract: Styrene is widely used in the production of various plastics, synthetic rubber and resins. Occupational exposure occurs mainly via inhalation and relatively high exposure occurs due to its use in manual application techniques. The aim of this study was to evaluate if SO-Hb adducts are a suitable biomarker for assessing occupational exposure to styrene. Seventy-five reinforced plastic workers and 77 control subjects were studied. In the selected population the main urinary styrene metabolites and the styrene oxide N-terminal valine (SO-Hb) adducts in human globin were quantified. The levels of SO-Hb adducts were significantly higher (p<0.01) in the exposed subjects (5.98pmol/g globin) when compared with controls (2.59pmol/g globin) and a significant difference was found in levels of SO-Hb adducts between non-smokers and smokers among the control group. From our data we conclude that SO-Hb adduct measurement is a sensitive and specific means of assessing exposure to styrene at the occupational and environmental level. [Copyright &y& Elsevier]

2,4,6-Trinitrotoluene (TNT) is an important occupational and environmental pollutant. In TNT-exposed humans, notable toxic manifestations have included aplastic anaemia, toxic hepatitis, cataracts, hepatomegaly and liver cancer. Therefore, it is important to develop protection measures and to monitor workers involved in the clean-up of ammunition sites. Haemoglobin (Hb) adducts of TNT, 4-amino-2,6-dinitrotoluene (4ADNT) and 2-amino-4,6-dinitrotoluene (2ADNT), and the urine metabolites of TNT, 4ADNT and 2ADNT were found in 22-50% of the exposed workers, but not in the control group. The exposed workers were wearing protective equipment. The levels of erythrocytes, haemoglobin, creatinine, serum glutamic pyruvic transaminase and lymphocyte levels were significantly lower in the exposed workers than in the non-exposed workers. The levels of blood urea and reticulocytes were significantly higher in the exposed workers than in the non-exposed workers. Headache (26%), mucous membrane irritation (16%), sick leave (18%), lassitude (8%), anxiety (6%), shortness of breath (3%), nausea (5%) and allergic reactions (8%) were reported by the exposed workers. In a further analysis the U-4ADNT levels and the Hb-adduct levels were compared to the blood parameter and the health effects. The blood parameters were not significantly different between the U-4ADNT positive and U-4ADNT-negative group. Headache, mucous membrane irritation, sick leave, lassitude, anxiety, shortness of breath and allergic reactions were statistically not different between the two groups. Also in the workers with Hb-4ADNT adducts no significant negative changes were seen in regards to the changes of the blood parameters or the health effects. According to the results of the present study, it appears that the blood parameter changes and the health effects are more influenced by other factors than by the internal exposure to TNT. [ABSTRACT FROM AUTHOR]

4,4'-Methylenediphenyl diisocyanate (MDI) is the most important isocyanate in the manufacture of polyurethanes, dyes, pigments and adhesives. High concentrations of isocyanates are a potent respiratory irritant. Therefore, it is important to develop methods to monitor exposure to such compounds. We monitored biological samples from 40 non-exposed and 45 exposed construction site workers. 4,4'-Methylenedianiline (MDA) and N-acetyl-4,4'-MDA (AcMDA) were determined from untreated urine (U-MDA, U-AcMDA) and MDA was analysed from acid-treated urine (U-MDA-tot). Haemoglobin (Hb) adducts of MDA (Hb-MDA) were determined in all workers. The levels of biomarkers decreased in the following order: U-MDA-tot>U-AcMDA>U-MDA>Hb-MDA. The same order was found for the percentage of samples, which were found positive in exposed workers: 100%, 91%, 91%, 27%. The urine levels U-MDA-tot correlate with U-MDA, U-AcMDA and Hb-MDA with r=0.79, 0.86 and 0.39, respectively (Spearman rank order, p<0.01). U-AcMDA correlates with U-MDA and Hb-MDA with r=0.77 and 0.47, respectively (p<0.01). U-MDA correlates with Hb-MDA (r=0.38, p<0.05). The levels in the controls were significantly lower than in the exposed workers for all compounds (Mann-Whitney test, p<0.01). The median isocyanate-specific IgE-level was higher in the exposed workers, but the difference was statistically not significant. The change of the biomarker levels was compared in a group of workers (n=20), which were analysed prior to isocyanate exposure and after the exposure for ∼4-7 months. All urine MDA metabolites and the Hb-adduct levels increased significantly (Wilcoxon sign test, p<0.01). Total IgE increased significantly after the exposure with isocyanate activity (p<0.01). With the present work it could be shown that outdoor workers are exposed to a similar extent as workers from a MDI factory. [ABSTRACT FROM AUTHOR]

Abstract: Styrene is one of the most important organic chemicals used worldwide. In humans, styrene metabolism involves oxidation by cytochrome P450 monooxygenases (CYPs) to styrene-7,8-oxide, an epoxide thought to be responsible for the genotoxic effects of styrene exposure, and detoxification by means of epoxide hydrolase (mEH) and glutathione S-transferases (GSTs). The objective of this study was to investigate if genetic polymorphisms of metabolic enzymes modulate the level of urinary styrene metabolites and styrene oxide adducts with N-terminal valine of human globin (SO-Hb) in 75 workers occupationally exposed to styrene and 77 unexposed controls. The mean air concentration of styrene in the breathing zone of workers (30.4ppm) was higher than the threshold limit value of 20ppm recommended by the American Conference of Governmental Industrial Hygienists (ACGIH), and the biological exposure index adopted by the ACGIH for exposure to styrene prior to the next shift (MA+PGA=400mg/g creatinine) was exceeded, indicating that styrene exposure for this group of workers was higher than recommended. A highly significant correlation was observed between styrene concentration in the breathing zone and the MA+PGA in urine of workers (r =0.85, P <0.001). The levels of SO-Hb adducts in exposed workers were significantly increased as compared with controls, although no difference was observed between subjects stratified as high and medium exposure categories based on MA+PGA excretion. Regarding the effect of the genetic polymorphisms we found that the level of SO-Hb adducts might be modulated by the predicted mEH enzymatic activity in the exposed workers. From our data we conclude that SO-Hb adduct measurement is a complementary method to MA+PG measurement for assessing exposure to styrene at occupational and environmental levels, which reflects a more extensive exposure period. [Copyright &y& Elsevier]

Abstract: Acrylamide is a food toxicant suspected to be carcinogenic to humans. It is formed in the heat processing of carbohydrate-rich food. A current issue in food safety is whether acrylamide actually represents a risk for human health. At present, available information is insufficient to reach any conclusions. Inter alias, a still unclear matter is the fraction of acrylamide ingested by food that is absorbed and metabolized. This study compared the in vivo relative absorption of acrylamide formed in cooked food with that of the pure compound dissolved in drinking water using the pig (25 Italian Large White females) as the animal model. Acrylamide intakes of about 0.8 and 8μgkg−1 pig body wtday−1 equal to one and ten times, respectively, the maximum average acrylamide daily intake for humans from the diet (expressed on a body wt basis) in industrialized countries, were chosen for the study. Adducts with the N-terminal valine of haemoglobin formed by acrylamide and its epoxide metabolite glycidamide, were used as exposure markers. Analyses were carried out by gas chromatography/mass spectrometry following in-house method validation. Both for the low and the high dose regimen, the glycidamide adduct levels in swine globins were lower of the limit of quantification of the method. As concerns acrylamide adducts, it was found that the relative absorption of acrylamide from feed and water was the same and that there is a direct proportionality between the adduct concentration and acrylamide intake. [Copyright &y& Elsevier]

For workers exposed to 4-chloronitrobenzene (4CNB), the major metabolites were determined. Urine were analysed before and after acid hydrolysis to qualify the free and conjugated metabolites of 4CNB. Three conjugated metabolites were identified in exposed workers: the mercapturic acid N-acetyl-S-(4-nitrophenyl)-L-cysteine (NANPC) was the only metabolite detected in non-hydrolysed urine, and accounted for approximately 51% of the total metabolites detected. The two remaining metabolites 4-chloroaniline (4CA) and 2-chloro-5-nitrophenol (CNP) were identified as cleavage products in hydrolysed urine, and accounted for approximately 18 and 30% of the total metabolites detected, respectively. No metabolites were found in factory controls within the limits of quantitation (LOQ) of the assay. There is a moderate correlation between NANPC and both 4CA and CNP. The correlation between 4CA and CNP is minor. The correlation between the total metabolites and both 4CA and CNP are good. The best correlation was found between the total metabolites and NANPC. There is a moderate inverse correlation between age and the creatinine levels. The raw metabolite levels CNP and NANPC decrease with age. 4CA, NANPC and the total metabolite levels correlate with the haemoglobin adduct levels . The urine metabolites increase and correlate significantly with the creatinine levels. NANPC is the most appropriate biomarker in the urine for a recent absorbed dose of 4CNB, since NANPC reflects the levels of 4CA and CNP and is the most prevalent metabolite detected in all the exposed workers. [ABSTRACT FROM AUTHOR]

Propylene oxide (PO) is an important industrial compound and a directly acting mutagen. Human exposure to PO can be monitored by the determination of haemoglobin adducts. An immunoassay that quantifies the N-terminal adduct N-(2-hydroxypropyl)valine in whole haemoglobin was developed and its potential usefulness as a tool for biologically monitoring occupational exposure was demonstrated. Analytical reliability was confirmed in a comparative study with GC-MS (range 3.7–992 nmol g -1 haemoglobin (Hb), correlation coefficient 0.99, n =10). The assay has been configured as a competitive enzyme-linked immunosorbent assay to facilitate the rapid throughput of samples. The assay employs a whole blood matrix and has a working range of 2–250 pmol g -1 Hb. It does not appear to be affected by structurally similar metabolites and has been used to determine adducts in human blood samples. The first results in potentially exposed workers indicate the assay's high potential usefulness in routine occupational biomonitoring of exposure to PO. [ABSTRACT FROM AUTHOR]

Abstract: Acrylamide (AA) is a food-borne toxicant suspected to be carcinogenic to humans. It is formed in the heating process of starch-containing food. Currently, there is a great discussion about the possible human health risks connected with the dietary uptake of acrylamide. Haemoglobin adducts of acrylamide and its oxidative metabolite glycidamide are both markers of biochemical effect. However, because glycidamide has a higher carcinogenic potency than acrylamide itself, the glycidamide adduct might mirror the genotoxicity better than acrylamide adducts. In order to gain more information about the human metabolism of acrylamide, we investigated a small group of persons for the effective internal doses of acrylamide and glycidamide using haemoglobin adducts as parameters of biochemical effect. The collective was subdivided into non-smokers (n=13) and smokers (n=16) by determining the smoking-specific acrylonitrile haemoglobin adduct (N-cyanoethylvaline, CEV). The mean values for the adducts of acrylamide (N-2-carbamoylethylvaline, AAVal) and glycidamide (N-(R,S)-2-hydroxy-2-carbamoylethylvaline, GAVal) in nonsmokers was 19 pmol/g globin AAVal (range 7 – 31 pmol/g globin) and 17 pmol/g globin GAVal (range 9 – 23 pmol/g globin). For smokers mean levels of AAVal were 80 pmol/g globin (range: 25 – 199 pmol/g globin) and those of GAVal were 53 pmol/g globin (range: 22 – 119 pmol/g globin). Metabolism to glycidamide turned out to be significantly more effective in non-smokers than in the higher exposed smokers. Compared with studies in rats, the metabolic conversion of acrylamide to glycidamide as measured by haemoglobin adducts seems to occur to a similar extent in humans as in rats. Risk estimations on acrylamide based on experimental data obtained in rats obviously did not overestimate the cancer risk for the general population. Furthermore, our results might indicate that the dose-response curve for acrylamide is not linear. This would be in line with the results of animal experiments on rodents. [Copyright &y& Elsevier]

Ethylene oxide (EO) is an important industrial compound and a directly acting mutagen. Human exposure to it can be monitored by the determination of haemoglobin (Hb) adducts. An immunoassay that quantifies the N-terminal adduct N-(2-hydroxyethyl)valine in whole blood was developed and its potential usefulness as a tool for biologically monitoring occupational exposure demonstrated. Analytical reliability was confirmed in a comparative study with gas chromatography-mass spectrometry (range 0.040-589 nmol g -1 Hb, correlation coefficient 0.98, n =10). The assay was configured as a competitive enzyme-linked immunosorbent assay to facilitate the rapid throughput of samples. The assay uses a whole blood matrix and has a working range of 10-10 000 pmol N-(2-hydroxethyl)valine g -1 Hb. The assay does not appear to be affected by structurally similar metabolites and has been used to determine adducts in human blood samples. The first results from potentially exposed workers indicate the assay might be a powerful tool for the routine occupational biomonitoring of EO exposure. [ABSTRACT FROM AUTHOR]

Objectives: Acrylamide (AA) (CAS No 79-06- 1) has most recently been identified as a food-borne toxicant generated during the heating process of starch-containing foods. It was the aim of the present study to investigate the trans-placental exposure of newborn infants to this possible human carcinogen by analysis of the specific haemoglobin adduct of AA (N- 2-carbamoylethylvaline, AAV) in the blood of mothers and the corresponding umbilical cord blood of neonates as a parameter of biochemical effects. Methods: We investigated the blood of 11 women advanced in pregnancy (one smoker, ten non-smokers) and the corresponding umbilical cord blood of neonates for the N-terminal haemoglobin adducts of AA (AAV) and the smoking-specific adduct of acrylonitrile (CAS No 107-13-1) (N-cyanoethylvaline, CEV). The limit of detection (LOD) was 5 pmol/g globin for AAV and 4 pmol/g globin for CEV. Results: AAV could be determined in all blood samples of the mothers (median 21 pmol/g globin, range 18-104 pmol/g globin) as well as in the umbilical cord blood of neonates (median 10 pmol/g globin, range 6-43 pmol/g globin). The highest values were detected in the blood of the smoking mother and her child. CEV was detected only in the blood of the smoking mother (185 pmol/g globin) and the corresponding umbilical cord blood (69 pmol/g globin). Discussion: AAV adduct levels in non-smoking mothers and neonates showed a good correlation (r = 0.859). The concentration of AA adducts in the blood of neonates is approximately 50% of the adduct level found in the blood of the mother. In view of the shorter life span of neonatal erythrocytes and the lower body weight of newborn infants, the relative internal dose of AA in neonates (in microgrammes per kilogramme body weight) must be assumed to be at least equal to that of the mother. Because of the high cell-replication rates during foetal development, trans-placental exposure of neonates to AA might raise concerns. Neonates of smoking mothers take up much higher doses of AA than those of non-smoking mothers. [ABSTRACT FROM AUTHOR]

Many industrial bulk chemicals are oxiranes or alkenes that are easily metabolised to oxiranes in mammalian systems. Many oxiranes may react with DNA and are therefore mutagenic in vitro. Some oxiranes have been shown to be carcinogenic in rodents in vivo as well. Despite the very limited evidence of the carcinogenicity of oxiranes in humans, they should be considered potential human carcinogens. As a consequence, exposure to these compounds should be minimised and controlled. Twenty-five years ago, Ehrenberg and co-workers suggested that exposure to oxiranes might be determined through the measurement of the adducts they form with haemoglobin (Hb). Ten years later, a modification of the Edman degradation was developed at Stockholm University that allowed determination of adducts with the N-terminal valine of Hb by GC–MS. In our laboratory, this methodology was modified and adapted for analysis on an industrial scale. Since 1987, exposure of operators in our facilities to ethylene oxide (EO) has been routinely monitored by determination of N-(2-hydroxyethyl)valine in Hb. Biological monitoring programmes for propylene oxide (PO) and 1,3-butadiene (BD) were developed later. In this review, the methodology and its results are discussed as a tool in human risk assessment of industrial chemicals. Two major advantages of Hb adduct determinations in risk assessment are (1) the qualitative information on the structure of reactive intermediates that may be obtained through the mass spectrometry, which may provide insight in the molecular toxicology of compounds such as BD, and (2) the possibility of reliable determination of exposure over periods of several months with limited number of samples for compounds such as ethylene oxide (EO), propylene oxide (PO) and BD which form stable adducts with Hb. Since good correlations between the airborne concentrations of these chemicals with their respective adducts have been established, Hb adducts can also be used to quantitate airborne exposure which is of paramount importance as exposure assessment is usually one of the weaker parameters in risk assessment. [ABSTRACT FROM AUTHOR]

Arylamines and nitroarenes are very important intermediates in the industrial manufacture of dyes, pesticides and plastics, and are significant environmental pollutants. The metabolic steps of N-oxidation and nitroreduction to yield N-hydroxyarylamines are crucial for the toxic properties of arylamines and nitroarenes. Nitroarenes are reduced by microorganisms in the gut or by nitroreductases and aldehyde dehydrogenase in hepatocytes to nitrosoarenes and N-hydroxyarylamines. N-Hydroxyarylamines can be further metabolized to N-sulphonyloxyarylamines, N-acetoxyarylamines or N-hydroxyarylamine N-glucuronide. These highly reactive intermediates are responsible for the genotoxic and cytotoxic effects of this class of compounds. N-Hydroxyarylamines can form adducts with DNA, tissue proteins, and the blood proteins albumin and haemoglobin in a dose-dependent manner. DNA and protein adducts have been used to biomonitor humans exposed to such compounds. All these steps are dependent on enzymes, which are present in polymorphic forms. This article reviews the metabolism of arylamines and nitroarenes and the biomonitoring studies performed in animals and humans exposed to these substances. [ABSTRACT FROM AUTHOR]

Objective. Based on our previous studies we analysed DNA and haemoglobin adducts, single-strand breaks in DNA, chromosomal aberrations and HPRT mutant frequencies in styrene-exposed workers, in relation to employment time, with respect to markers of individual susceptibility. In our study the risk assessment of genotoxic styrene in occupationally exposed humans is reviewed in the light of adaptation and/or population selection. Various styrene-induced DNA adducts (representing important integral measures of DNA damage) are discussed as potential powerful biomarkers of styrene exposure, with respect to DNA repair. Methods. The complexity of followed markers required multiple methodological approaches, including modified 32P-post-labelling assay, PCR-based methods for genotyping, cytogenetic analysis, T-cell cloning assay, comet assay and tests for DNA repair capacity, and statistical tests. Results. Haemoglobin and DNA adducts, single-strand breaks in DNA, chromosomal aberrations and mutant frequencies at the HPRT gene were assessed with respect to the duration of exposure. No apparent accumulation of these biomarkers was found to be dependent on the years of employment, suggesting that adaptive processes play a role in chronically exposed workers. Statistically significant differences between the exposed and control individuals were found for the deduced epoxide hydrolase activity (P=0.041) and the frequency of heterozygous Ala/Val genotype in GSTP1, exon 5 (P=0.025). Significantly increased DNA repair capacity was found in styrene-exposed laminators compared with control individuals. Conclusions. In the light of present knowledge well-designed population studies on styrene genotoxicity are needed. Such studies should cover important areas of individual susceptibility (metabolising and repair enzymes, distributions of genetic polymorphisms) and possible role of adaptation, as well as the crucial role of DNA repair. Additionally, genotoxic effects of in-vivo formed 3,4-styrene oxide should be also addressed. [ABSTRACT FROM AUTHOR]

Case reports of human accidental poisonings point to significant individual differences in human acrylonitrile metabolism and toxicity. A cohort of 59 persons with industrial handling of low levels of acrylonitrile has repetitively been studied from 1994 through 1999 as part of a medical surveillance programme. The analyses included adduct determinations of N-terminal N-(cyanoethyl)valine in haemoglobin and genotypings of the following cytochrome P-450 2E1 (CYP2E1) polymorphisms: G−1259C and C−1019T (two subjects heterozygous), A−316G (three subjects heterozygous), T−297A (15 subjects heterozygous), G−35T (eight subjects heterozygous), G4804A (two subjects heterozygous), T7668A (six subjects heterozygous). N-(Cyanoethyl)valine adduct levels were, if any, only slightly influenced by smoking and mainly determined by the external acrylonitrile exposures. The individual means and medians of N-(cyanoethyl)valine levels over the entire observation period were compared with the CYP2E1 variants (Wilcoxon rank sum test). No influences of the investigated CYP2E1 polymorphisms on the N-(cyanoethyl)valine levels appeared at the 5% level. However, there was a trend, at a level of P&ap;0.1, pointing to higher acrylonitrile-specific adduct levels in persons with the A−316G mutation. Higher adduct levels would be compatible with a slower CYP2E1-mediated metabolism of acrylonitrile and with lower extents of toxification to cyanide. [Copyright &y& Elsevier]

The high priority of monitoring workers exposed to nitrobenzene is a consequence of clear findings of experimental carcinogenicity of nitrobenzene and the associated evaluations by the International Agency for Research on Cancer. Eighty male employees of a nitrobenzene reduction plant, with potential skin contact with nitrobenzene and aniline, participated in a current medical surveillance programme. Blood samples were routinely taken and analysed for aniline, 4-aminodiphenyl (4-ADP) and benzidine adducts of haemoglobin (Hb) and human serum albumin (HSA). Also, levels of methaemoglobin (Met-Hb) and of carbon monoxide haemoglobin (CO-Hb) were monitored. Effects of smoking were straightforward. Using the rank sum test of Wilcoxon, we found that very clear-cut and statistically significant smoking effects (about 3-fold increases) were apparent on CO-Hb (P=0.00085) and on the Hb adduct of 4-ADP (P=0.0006). The mean aniline-Hb adduct level in smokers was 1.5 times higher than in non-smokers; the significance (P=0.05375) was close to the 5% level. The strongest correlation was evident between the Hb and HSA adducts of aniline (rs=0.846). Less pronounced correlations (but with P values < 0.02) appeared between aniline-Hb and 4-ADP-Hb adducts (rs=0.388), between 4-ADP and 4-ADP-HSA adducts (rs=0.373), and between 4-ADP-Hb and aniline-HSA adducts (rs=0.275). In view of the proposal for additional use of the aniline-HSA adduct for biological monitoring, particularly in cases of acute overexposures or poisonings, the strong correlation of the Hb and HSA conjugates is noteworthy; the ratio aniline-HSA:aniline-Hb was 1:42 for the entire cohort. [ABSTRACT FROM AUTHOR]

Objective: The influence of area of residence on haemoglobin (Hb) adducts of 4-aminobiphenyl (4-ABP), o-, m-, p-toluidine and o-anisidine was investigated in children from three different-sized Bavarian cities – Munich, Augsburg and Eichstätt, with 1,300,000, 250,000 and 13,000 inhabitants, respectively – and was compared with that of exposure to environmental tobacco smoke (ETS). Methods: Blood samples from Munich (n=34) and Eichstätt (n=64) were from children attending the Paediatric Clinic of the Technical University of Munich (TUM) or a practice in Eichstätt, respectively. Blood samples (n=126) together with urine samples (n=88) were collected from Augsburg children during school medical examination. Personal data including possible sources of ETS exposure were obtained at the interview. Hb adduct levels were analysed by a gas chromatographic method, using mass spectrometry with selected-ion monitoring. Urinary cotinine was determined by radioimmunoassay. Results: 4-ABP Hb adduct levels in children from Munich were 1.5 and 1.2 times higher than those in children from Eichstätt and Augsburg (P<0.001). Children from Munich also had significantly higher Hb adduct levels of monocyclic aromatic amines than did children from Eichstätt and, except for o-toluidine, children from Augsburg (P<0.005). Compared with children from Eichstätt, children from Augsburg had higher Hb adduct levels of 4-ABP, o- and m-toluidine (P<0.01) but not p-toluidine and o-anisidine. In a multivariate analysis, gender, age and body mass index had no consistent influence on Hb adducts. ETS exposure resulted in a slight, nonsignificant increase in 4-ABP Hb adduct levels. In contrast, adduct levels from monocyclic aromatic amines were consistently decreased in ETS-exposed children (significant for o- and m-toluidine, P<0.05). Conclusions: Hb adducts from aromatic amines in children were strongly influenced by site of residence, whereas ETS exposure did not significantly increase the adduct levels. [ABSTRACT FROM AUTHOR]

According to the kinetic theory for the build-up and elimination of haemoglobin (Hb) adducts, unstable Hb adducts are simultaneously eliminated by zero-order Hb turnover and first-order chemical instability. Thus, the elimination of unstable Hb adducts is non-linear with respect to time. Nonetheless, many studies of Hb adduct stability have characterized the elimination of Hb adducts using linear zero-order or linear first-order models. This paper demonstrates the use of non-linear regression to estimate the first-order rate constant of Hb adduct instability (k) using data on the elimination of Hb adducts in rats dosed with benzene or ortho -toluidine. Results obtained using non-linear regression models are compared with results from the more commonly employed zero- and first-order linear models. It is shown that exposure estimates based on measured levels of unstable Hb adducts can be severely biased if zero-order turnover is assumed. Furthermore, based on published data, estimates of k are subject to estimated relative biases in the range of -4% to 96% when first-order linear models are used to characterize Hb adduct instability. [ABSTRACT FROM AUTHOR]

In view of the established extrapulmonary cancer sites targeted by smoking a multiplicity of compounds and mechanisms might be involved. It has been debated that smoking caused increased incidence of N-methylvaline at the N-terminus of haemoglobin. Because this could indicate a relevance of methylating nitrosamines in tobacco smoke, data are presented from an industrial cohort of 35 smokers and 21 non-smokers repeatedly monitored between 1994 and 1999. In general, N-methylvaline adduct levels in haemoglobin of smokers were approximately 50% higher than those of non-smokers. The smoking-induced methylation of haemoglobin is likely to be caused by dimethylnitrosamine (N-nitroso-dimethylamine), a major nitrosamine in side-stream tobacco smoke. The biomonitoring data emphasise the potential value of N-methylvaline as a smoking-related biomarker and call for intensified research on tobacco smoke compounds that lead to macromolecular methylation processes. [ABSTRACT FROM AUTHOR]

High levels of haemoglobin (Hb) adducts from 4-aminobiphenyl (4-ABP), a proven human carcinogen, have been reported in untreated animals from different laboratories fed various commercial standard diets. Therefore, the impact of dietary modifications on 4-ABP Hb adducts was investigated. Female Sprague-Dawley rats were fed a regular standard diet or three different test diets for 4 weeks. 4-ABP Hb adducts were significantly lower in rats on vegetable-based test diets #2 (596±183 pg/g Hb, P=0.028) and #3 (537±48 pg/g Hb, P=0.009) compared with controls (974±154 pg/g Hb). Cereal-based test diet #1 (1080±388 pg/g Hb) had no influence on the basal Hb adduct levels determined before the start of the experiment (1054±163 pg/g Hb). In conclusion, the body burden of rats with 4-ABP could be significantly reduced by dietary modifications. [ABSTRACT FROM AUTHOR]

Aromatic amines are important intermediates in industrial manufacturing. They are used in a large number of products, such as pesticides, dyes, plastics and pharmaceuticals. The parent arylamines can be metabolically released from these arylamine-based compounds and form DNA and protein adducts after N-oxidation to N-hydroxy arylamines. Aromatic amine derivatives, including the industrial intermediates acetoacetanilide, acetoacet-m-xylidide and N-ethylaniline, were examined for their ability to form Hb adducts in rats as potential biomarkers of exposure. The haemoglobin binding indices (HBI=binding [mmol mol-1Hb]/dose [mmol kg-1 body weight]) of the arylamines were determined 24 h after oral administration to female Wistar rats. The precipitated haemoglobin was dissolved in 0.1 M sodium hydroxide in the presence of internal standards. After hexane extraction the released arylamines were analysed by gas chromatography-mass spectrometry (GC-MS). For aniline released from acetoacetanilide an HBI of 15 and for 2,4-dimethylaniline released from acetoacet-m-xylidide an HBI of 0.129 were determined. The HBIof aniline released from N-ethylaniline was 45. [ABSTRACT FROM AUTHOR]

Fifty-nine persons with industrial handling of low levels of acrylonitrile (AN) were studied. As part of a medical surveillance programme an extended haemoglobin adduct monitoring [N-(cyanoethyl)valine, CEV; N-(methyl)valine, MV; N-(hydroxyethyl)valine, HEV] was performed. Moreover, the genetic states of the polymorphic glutathione transferases GSTM1 and GSTT1 were assayed by polymerase chain reaction (PCR). Repetitive analyses of CEV and MV in subsequent years resulted in comparable values (means, 59.8 and 70.3 μg CEV/l blood; 6.7 and 6.7 μg MV/l blood). Hence, the industrial AN exposures were well below current official standards. Monitoring the haemoglobin adduct CEV appears as a suitable means of biomonitoring and medical surveillance under such exposure conditions. There was also no apparent correlation between the CEV and HEV or CEV and MV adduct levels. The MV and HEV values observed represented background levels, which apparently are not related to any occupational chemical exposure. There was no consistent effect of the genetic GSTM1 or GSTT1 state on CEV adduct levels induced by acrylonitrile exposure. Therefore, neither GSTM1 nor GSTT1 appears as a major AN metabolizing isoenzyme in humans. The low and physiological background levels of MV were also not influenced by the genetic GSTM1 state, but the MV adduct levels tended to be higher in GSTT1− individuals compared to GSTT1+ persons. With respect to the background levels of HEV adducts observed, there was no major influence of the GSTM1 state, but GST− individuals displayed adduct levels that were about 1/3 higher than those of GSTT1+ individuals. The coincidence with known differences in rates of background sister chromatid exchange between GSTT1− and GSTT1+ persons suggests that the lower ethylene oxide (EO) detoxification rate in GSTT1− persons, indicated by elevated blood protein hydroxyethyl adduct levels, leads to an increased genotoxic effect of the physiological EO background. [ABSTRACT FROM AUTHOR]

Propanil is one of the major herbicides used on rice-paddies and is thought to produce adverse health effects through the action of its metabolite 3,4-dichloroaniline (3,4-DCA). T he feasibility of monitoring human exposure to propanil on the basis of 3,4-DCA adducts to haemoglobin (Hb) was investigated. We developed a method based on gas chromatography negative ion chemical ionization-mass spectrometry (NICI-GC-MS) to quantify 3,4-DCA released from human Hb after alkaline hydrolysis of the protein. 3,4-DCA-Hb adducts were identified in agricultural workers exposed to propanil and were detectable even 4 months after the last herbicide application. Urine samples collected at the same time had no measurable level of 3,4-DCA. 3,4-DCA-Hb adducts might be useful for monitoring human exposure to 3,4-DCA from agricultural sources. [ABSTRACT FROM AUTHOR]

The aim of this study was to determine whether haemoglobin adducts Hb of hexahydrophthalic anhydride HHPA and HHPA specific immunoglobulin G IgG can be used as biomarkers of exposure to HHPA. The exposures of HHPA in 10 workers were determined from the mean urinary hexahydrophthalic acid HHP acid levels range 76-3300 nmol HHP acid mmol-1 creatinine during a period of 4 weeks. Blood was collected at the end of the period and Hb-HHPA adducts were analysed by gas chromatography mass spectrometry. The Hb-HHPA adduct levels ranged from 0.45 to 24.7 pmol g-1 Hb. There was a close correlation between the urinary HHP acid levels and the amount of Hb-HHPA adducts r = 0.87 . One day exposures to HHPA and methylhexahydrophthalic anhydride MHHPA in 142 workers were determined from analysis of urinary HHP acid range 0-3300 nmol HHP acid mmol-1 creatinine and methylhexahydrophthalic acid MHHP acid; range 0-1700 nmol MHHP acid mmol-1 creatinine . HHPA specific IgG were analysed in the 142 workers with an ELISA method. The optical density for HHPA specific IgG varied between 0 and 1.25. There was no statistically significant correlation between the sum of the urinary HHP acid and MHHP acid and the HHPA specific IgG r = 0.12; p = 0.14 . Thus, Hb-HHPA adducts seem to be applicable as biomarkers of exposure to HHPA while the possible role of HHPA specific IgG as an indicator of exposure has to be further evaluated. [ABSTRACT FROM AUTHOR]

Aromatic amines are metabolically activated by N-oxidation of either the amine or the acetamide as a first step and esterification of the resulting N-hydroxyl derivatives as a second step. Both pathways may lead to DNA-adducts and subsequently to DNA lesions and mutations. Since the accumulation of non-acetylated adducts has been associated with tumour initiating properties, the balance between acetylation and deacetylation may greatly influence the biological effect. Hydrolysable haemoglobin adducts representing the bioavailability of N-hydroxylamines and the corresponding nitroso-derivatives were analysed following oral administration to female Wistar rats of two arylamine-acetamide couples: 4-aminobiphenyl and 2-aminofluorene, and two arylamine-acetamide-diacetamide triples: benzidine and 3,3′-dichlorobenzidine. The results show that the mono-acetamides are readily deacetylated in vivo whereas the diacetamides are not. A dynamic equilibrium is indicated to exist between acetylation and deacetylation, which depends on substrate specificity, and the role of deacetylation is emphasised. In addition, acetylation polymorphism was studied with 4-chloroaniline and 3,3′-dichlorobenzidine in slow acetylating A/J and rapid acetylating C57BL/6J mice. The slow acetylator genotype was associated with significantly higher haemoglobin-adduct levels for both arylamines. The results provide additional support for the use of haemoglobin adducts in biomonitoring as a dosimeter for the biologically active dose of arylamines/arylacetamides. Moreover, biomonitoring of haemoglobin adducts may provide information about an individual's susceptibility to the toxic and carcinogenic effects of these chemicals. [ABSTRACT FROM AUTHOR]

N,N-dimethylformamide (DMF) is a commonly used industrial solvent. The formation of some metabolites of DMF in humans occurs via N-methylcarbamoylated species (e.g. N-methylcarbamoylated glutathione). The aim of our study was to investigate whether DMF leads to N-methylcarbamoylated adducts at the N-terminal valine of haemoglobin (Hb). Therefore, Hb adduct levels of ten DMF exposed workers and ten controls were analysed by a specific and sensitive detection method using capillary gas chromatography and a mass selective detector (GC/MS). Using this method we were able to show for the first time that Hb adducts are formed during the metabolism of DMF in humans. The general population, however, shows still unidentified background levels of this adduct which are on average lower by a factor of 50. The pathway for the formation of the investigated DMF-Hb adduct in workers exposed to DMF is still unknown. As identical adducts were also found after exposure to methylisocyanate (MIC), our work indicates the formation of MIC during the metabolism of DMF. The formation of Hb adducts with DMF and its relevance for occupational health is a subject of further research. [ABSTRACT FROM AUTHOR]