title: Quantitation of 4,4'-methylene diphenyl diisocyanate human serum albumin adducts.
abstract: 4,4'-Methylene diphenyl diisocyanate (herein 4,4'-MDI) is used in the production of polyurethane foams, elastomers, coatings, adhesives and the like for a wide range of commercial products. Occupational exposure to MDI levels above current airborne exposure limits can elicit immune mediated hypersensitivity reactions such as occupational asthma in sensitive individuals. To accurately determine exposure, there has been increasing interest in developing analytical methods to measure internal biomarkers of exposure to MDI. Previous investigators have reported methodologies for measuring MDI diamine metabolites and MDI-Lysine (4,4'-MDI-Lys) adducts. The purpose of this study was to develop and validate an ultra performance liquid chromatography isotope dilution tandem mass spectrometry (UPLC-ID/MS/MS) quantitation methodviaa signature peptide approach to enable biomonitoring of 4,4'-MDI adducted to human serum albumin (HSA) in plasma. A murine, anti-4,4'-MDI monoclonal IgM antibody was bound to magnetic beads and utilized for enrichment of the MDI adducted HSA. Following enrichment, trypsin digestion was performed to generate the expected 414 site (primary site of adduction) 4,4'-MDI-adducted HSA signature peptide that was quantified by UPLC-ID/MS/MS. An Agilent 6530 UPLC/quadrupole time of flight MS (QTOF) system was utilized for intact adducted protein analysis and an Agilent 6490 UPLC/MS/MS system operated in multiple reaction monitoring (MRM) mode was utilized for quantification of the adducted signature peptide biomarker both forin chemicoand worker serum samples. Worker serum samples were initially screened utilizing the previously developed 4,4'-MDI-Lys amino acid method and results showed that 12 samples were identified as quantifiable for 4,4'-MDI-Lys adducts. The signature peptide adduct approach was applied to the 12 worker samples identified as quantifiable for 4,4'-MDI-Lys adducts. Results indicated no positive results were obtained above the quantification limit by the signature peptide approach. If the 414 site of lysine adduction accounted for 100% of the 4,4'-MDI adductions in the signature peptide adduct approach, the three highest quantifiable samples by the 4,4'-MDI-Lys method should have at least been detectable by the signature peptide method. Results show that although the 4,4'-MDI signature peptide approach is more selective, it is 18 times less sensitive than the 4,4'-MDI-Lys method, thus limiting the ability to detect adduct levels relative to the 4,4'-MDI-Lys amino acid method.
authors.affiliation: Toxicology, Environmental Research and Consulting, The Dow Chemical Company, United States.
authors.forename: Leah G
authors.initials: LG
authors.lastname: Luna
authors.affiliation: Allergy and Clinical Immunology Branch, Health Effects Laboratory Division, National Institute for Occupational Safety and Health, Centers for Disease Control and Prevention, United States.
authors.forename: Brett J
authors.initials: BJ
authors.lastname: Green
authors.affiliation: Toxicology, Environmental Research and Consulting, The Dow Chemical Company, United States.
authors.forename: Fagen
authors.initials: F
authors.lastname: Zhang
authors.affiliation: Toxicology, Environmental Research and Consulting, The Dow Chemical Company, United States.
authors.forename: Scott M
authors.initials: SM
authors.lastname: Arnold
authors.affiliation: Allergy and Clinical Immunology Branch, Health Effects Laboratory Division, National Institute for Occupational Safety and Health, Centers for Disease Control and Prevention, United States.
authors.forename: Paul D
authors.initials: PD
authors.lastname: Siegel
authors.affiliation: Toxicology, Environmental Research and Consulting, The Dow Chemical Company, United States.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Bartels
keywords: Adduct; Biomarkers; Diisocyanates; MDI; Mass spectrometry