Chemicals are measured regularly in air, food, the environment, and the workplace. Biomonitoring of chemicals in biological fluids is a tool to determine the individual exposure. Blood protein ...adducts of xenobiotics are a marker of both exposure and the biologically effective dose. Urinary metabolites and blood metabolites are short term exposure markers. Stable hemoglobin adducts are exposure markers of up to 120 days. Blood protein adducts are formed with many xenobiotics at different sites of the blood proteins. Newer methods apply the techniques developed in the field of proteomics. Larger adducted peptides with 20 amino acids are used for quantitation. Unfortunately, at present the methods do not reach the limits of detection obtained with the methods looking at single amino acid adducts or at chemically cleaved adducts. Therefore, to progress in the field new approaches are needed.
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•The EU Horizon 2020 project HBM4EU targets seven substance groups and two metals.•Best suitable exposure biomarkers, matrices and analytical methods were suggested.•The selection was ...based on generally applicable criteria, e.g. method sensitivity.•Some conflicts were identified between the criteria and common exposure biomarkers.•Stringent quality assurance and control measures should be included and reported.
The major purpose of human biomonitoring is the mapping and assessment of human exposure to chemicals. The European initiative HBM4EU has prioritized seven substance groups and two metals relevant for human exposure: Phthalates and substitutes (1,2-cyclohexane dicarboxylic acid diisononyl ester, DINCH), bisphenols, per- and polyfluoroalkyl substances (PFASs), halogenated and organophosphorous flame retardants (HFRs and OPFRs), polycyclic aromatic hydrocarbons (PAHs), arylamines, cadmium and chromium. As a first step towards comparable European-wide data, the most suitable biomarkers, human matrices and analytical methods for each substance group or metal were selected from the scientific literature, based on a set of selection criteria. The biomarkers included parent compounds of PFASs and HFRs in serum, of bisphenols and arylamines in urine, metabolites of phthalates, DINCH, OPFRs and PAHs in urine as well as metals in blood and urine, with a preference to measure Cr in erythrocytes representing Cr (VI) exposure. High performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) was the method of choice for bisphenols, PFASs, the HFR hexabromocyclododecane (HBCDD), phenolic HFRs as well as the metabolites of phthalates, DINCH, OPFRs and PAHs in urine. Gas chromatographic (GC) methods were selected for the remaining compounds, e.g. GC-low resolution MS with electron capture negative ionization (ECNI) for HFRs. Both GC–MS and LC-MS/MS were suitable for arylamines. New developments towards increased applications of GC–MS/MS may offer alternatives to GC–MS or LC-MS/MS approaches, e.g. for bisphenols. The metals were best determined by inductively coupled plasma (ICP)-MS, with the particular challenge of avoiding interferences in the Cd determination in urine. The evaluation process revealed research needs towards higher sensitivity and non-invasive sampling as well as a need for more stringent quality assurance/quality control applications and assessments.
Humans are potentially exposed to a large amount of chemicals present in the environment and in the workplace. In the European Human Biomonitoring initiative (Human Biomonitoring for the European ...Union = HBM4EU), acrylamide, mycotoxins (aflatoxin B1, deoxynivalenol, fumonisin B1), diisocyanates (4,4′-methylenediphenyl diisocyanate, 2,4- and 2,6-toluene diisocyanate), and pyrethroids were included among the prioritized chemicals of concern for human health. For the present literature review, the analytical methods used in worldwide biomonitoring studies for these compounds were collected and presented in comprehensive tables, including the following parameter: determined biomarker, matrix, sample amount, work-up procedure, available laboratory quality assurance and quality assessment information, analytical techniques, and limit of detection. Based on the data presented in these tables, the most suitable methods were recommended. According to the paradigm of biomonitoring, the information about two different biomarkers of exposure was evaluated: a) internal dose = parent compounds and metabolites in urine and blood; and b) the biologically effective = dose measured as blood protein adducts. Urine was the preferred matrix used for deoxynivalenol, fumonisin B1, and pyrethroids (biomarkers of internal dose). Markers of the biological effective dose were determined as hemoglobin adducts for diisocyanates and acrylamide, and as serum-albumin-adducts of aflatoxin B1 and diisocyanates. The analyses and quantitation of the protein adducts in blood or the metabolites in urine were mostly performed with LC-MS/MS or GC-MS in the presence of isotope-labeled internal standards. This review also addresses the critical aspects of the application, use and selection of biomarkers. For future biomonitoring studies, a more comprehensive approach is discussed to broaden the selection of compounds.
Isothiocyanates (ITCs) found in cruciferous vegetables have demonstrated cancer preventive activity in animals, and increased dietary intake of ITCs has been shown to be associated with a reduced ...cancer risk in humans. ITCs exert their cancer chemopreventive action by multiple mechanisms, for example, by modulating the activities of phase I and phase II drug metabolism enzymes, by inhibiting the cell cycle and histone deacetylase, and by causing apoptotic cell death. In cells, protein adducts account for most of total cellular ITC uptake at 4 h after treatment. The time course of this protein binding correlates well with the inhibition of proliferation and the induction of apoptosis. Animal studies have shown that glutathione conjugates are the major products of ITCs. The major urinary excretion products of ITCs in human are N-acetyl cysteine conjugates. Urinary metabolites might provide the exposure history of the last 24 h, if the urine of the full next day is collected. However, this is not feasible in large epidemiological studies. Furthermore, the mercapturic acids of ITC are not stable. Therefore, stable biomarkers are needed that reflect a larger time span of the ITC exposure history. We developed a method to determine stable (not cysteine adducts) reaction products of ITCs with albumin and hemoglobin in humans and mice. We reacted albumin with the ITCs: benzyl isothiocyanate (BITC), phenylethyl isothiocyanate (PEITC), sulforaphane (SFN), and allyl isothiocyanate (AITC). After enzymatic digestion, we found one major product with lysine using LC-MS/MS. The identity of the adducts was confirmed by comparing the analyses with synthetic standards: N 6-(benzylamino)carbonothioyllysine (BITC-Lys), N 6-{(2-phenylethyl)aminocarbonothioyl}lysine (PEITC-Lys), N 6-({3-(methylsulfinyl)propylamino}carbonothioyl)lysine (SFN-Lys), and N 6-(allylaminocarbonothioyllysine (AITC-Lys). The adduct levels were quantified by isotope dilution mass spectrometry using the corresponding new ITC-13C6 15N2lysines as internal standards. The applicability of the method was tested for biological samples obtained from different experiments. In humans consuming garden cress, watercress, and broccoli and/or in mice exposed chronically to N-acetyl-S-{(2-phenylethyl)aminocarbonothioyl}-l-cysteine, albumin and hemoglobin adducts were found. BITC-Lys, PEITC-Lys, and SFN-Lys released after enzymatic digestion of the proteins were quantified with LC-MS/MS. This new method will enable quantification of ITC adducts in blood proteins from large prospective studies about diet and cancer. Protein adducts are involved in the chemopreventive effects of ITCs. Therefore, blood protein adducts are a potential surrogate marker for the effects of ITCs at the cellular level. This new technique will improve the assessment of ITC exposure and the power of studies on the relationship between ITC intake and cancer.
Arylamines and nitroarenes are intermediates in the production of pharmaceuticals, dyes, pesticides, and plastics and are important environmental and occupational pollutants. N-Hydroxyarylamines are ...the toxic common intermediates of arylamines and nitroarenes. N-Hydroxyarylamines and their derivatives can form adducts with hemoglobin (Hb-adducts), albumin, DNA, and tissue proteins in a dose-dependent manner. Most of the arylamine Hb-adducts are labile and undergo hydrolysis in vitro, by mild acid or base, to form the arylamines. According to current knowledge of arylamine adduct-formation, the hydrolyzable fraction is derived from the reaction products of the arylnitroso derivatives that yield arylsulfinamide adducts with cysteine. Hb-adducts are markers for the bioavailability of N-hydroxyarylamines. Hb-adducts of arylamines and nitroarenes have been used for many biomonitoring studies for over 30 years. Hb-adducts reflect the exposure history of the last four months. Biomonitoring of urinary metabolites is a less invasive process than biomonitoring blood protein adducts, and urinary metabolites have served as short-lived biomarkers of exposure to these hazardous chemicals. However, in case of intermittent exposure, urinary metabolites may not be detected, and subjects may be misclassified as nonexposed. Arylamines and nitroarenes and/or their metabolites have been measured in urine, especially to monitor the exposure of workers. This review summarizes the results of human biomonitoring studies involving urinary metabolites and Hb-adducts of arylamines and nitroarenes. In addition, studies about the relationship between Hb-adducts and diseases are summarized.
1,5-Naphthalene diisocyanate (NDI) is used in the plastic industry as a curing agent. 1,5-Naphthalene diisocyanate is classified as a sensitizing agent. The objective of this study has been to ...develop biomonitoring methods for the evaluation of exposure to NDI.
We obtained blood and urine samples from a group of 20 male workers exposed to NDI. The workers answered a questionnaire about their exposure history, job description, the number of years with the company and the time spent working with NDI over the 10 days of the study. Total plasma, albumin, and urine were analyzed for the presence of 1,5-naphthalenediamine (NDA) after acid hydrolysis using gas chromatography-mass spectrometry (GC-MS).
1,5-Naphthalenediamine was found in about 60% of the samples obtained from the workers. 1,5-Naphthalenediamine was obtained after acid hydrolysis of plasma, albumin, and urine at levels up to 1.5 pmol NDA/mg of plasma proteins, 1.15 pmol NDA/mg of albumin, and 55.3 pmol NDA/ml of urine, respectively.
1,5-Naphthalenediamine found in urine correlates best with the plasma levels (r = 0.91, p < 0.01). The albumin-adduct levels did not correlate with the NDI-specific immunoglobulin E (IgE) or total IgE present in the workers. The adduct and metabolite levels correlate with the air levels of NDI. Int J Occup Med Environ Health 2017;30(4):579-591.
4,4′-Methylenediphenyl diisocyanate (MDI) is the most important of the isocyanates used as intermediates in the chemical industry. Among the main types of damage after exposure to low levels of MDI ...are lung sensitization and asthma. Albumin adducts of MDI might be involved in the etiology of sensitization reactions. This work presents a liquid chromatography (LC)-mass spectrometry (MS/MS) procedure for determination of isocyanate-specific albumin adducts in humans. MDI formed adducts with lysine of albumin: MDI-Lys and AcMDI-Lys. The MDI-Lys levels, 25th, 50th, 75th, 90th percentile, were 0, 65.2, 134, 244 fmol mg−1 and 0, 30.5, 57.4, 95.8 fmol mg−1 in the exposed construction and factory workers, respectively. This new biomonitoring procedure will allow assessment of suspected exposure sources and may contribute to the identification of individuals who are particularly vulnerable for developing bronchial asthma and other respiratory diseases after exposure to isocyanates.
Serum albumin (Alb) is the most abundant protein in blood plasma. Alb reacts with many carcinogens and/or their electrophilic metabolites. Studies conducted over 20 years ago showed that Alb forms ...adducts with the human carcinogens aflatoxin B1 and benzene, which were successfully used as biomarkers in molecular epidemiology studies designed to address the role of these chemicals in cancer risk. Alb forms adducts with many therapeutic drugs or their reactive metabolites such as β-lactam antibiotics, acetylsalicylic acid, acetaminophen, nonsteroidal anti-inflammatory drugs, chemotherapeutic agents, and antiretroviral therapy drugs. The identification and characterization of the adduct structures formed with Alb have served to understand the generation of reactive metabolites and to predict idiosyncratic drug reactions and toxicities. The reaction of candidate drugs with Alb is now exploited as part of the battery of screening tools to assess the potential toxicities of drugs. The use of gas chromatography-mass spectrometry, liquid chromatography, or liquid chromatography-mass spectrometry (LC-MS) enabled the identification and quantification of multiple types of Alb xenobiotic adducts in animals and humans during the past three decades. In this perspective, we highlight the history of Alb as a target protein for adduction to environmental and dietary genotoxicants, pesticides, and herbicides, common classes of medicinal drugs, and endogenous electrophiles, and the emerging analytical mass spectrometry technologies to identify Alb-toxicant adducts in humans.
Isocyanates such as 1,6-hexamethylene diisocyanate (HDI), 4,4′-methylenediphenyl diisocyanate, and toluene diisocyanate are highly reactive compounds that have a variety of commercial applications, ...including manufacturing polyurethane foam, elastomers, paints, adhesives, coatings, insecticides, and many other products. Their primary route of occupational exposure is through inhalation. Due to their high chemical reactivity, they are toxic and have adverse effects at the cellular and subcellular levels, leading to irritative and immunological reactions associated with lung disease. High concentrations of isocyanates are strong respiratory irritants. Bronchial sensitization and asthma are among the major adverse clinical reactions associated with low-level chronic exposure to isocyanates. Albumin adducts have been linked to the mechanism of occupational asthma caused by isocyanates. Isocyanates react in vivo with albumin, which is recognized by the immune system. Albumin adducts of isocyanates trigger immune responses and are probably the antigenic basis for isocyanate asthma. Sensitization to isocyanates is the main pathway for adverse health effects. Therefore, markers for the biologically effective dose such as albumin adducts of HDI are needed. A new isocyanate adduct of HDI with lysineN ε-(6-amino-hexyl-amino)carbonyl-lysine (HDI-Lys)was synthesized and characterized by 1H-NMR, 13C-NMR, and mass spectrometry (MS). Appropriate internal standardsHDI-Lys-4,4′-5,5′-d 4 (HDI-d 4-Lys) and N ε-(7-amino-heptyl-amino)carbonyl-lysine (Hep-Lys)were synthesized to establish a LC–MS/MS method for the analysis of HDI adducts in in vitro modified albumin and in workers. The presence of HDI-Lys was found after pronase digestion of albumin and confirmed by two independent chromatographic approaches: with a C8 reversed-phase column and with a hydrophilic interaction liquid chromatography column. Quantification was performed with positive electrospray ionization (ESI)–MS. The adduct peak found in vivo was confirmed with the less sensitive negative ESI–MS. In summary, these are new compounds and methods to determine isocyanate-specific adducts with albumin in workers exposed to HDI.
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