title: Quantitation of ortho-cresyl phosphate adducts to butyrylcholinesterase in human serum by immunomagnetic-UHPLC-MS/MS.
abstract: Tri-ortho-cresyl phosphate (ToCP) is an anti-wear, flame retardant additive used in industrial lubricants, hydraulic fluids and gasoline. The neurotoxic effects of ToCP arise from the liver-activated metabolite 2-(o-cresyl)-4H-1,3,2-benzodioxaphosphoran-2-one (cresyl saligenin phosphate or CBDP), which inhibits esterase enzymes including butyrylcholinesterase (BChE). Following BChE adduction, CBDP undergoes hydrolysis to form the aged adduct ortho-cresyl phosphoserine (oCP-BChE), thus providing a biomarker of CBDP exposure. Previous studies have identified ToCP in aircraft cabin and cockpit air, but assessing human exposure has been hampered by the lack of a laboratory assay to confirm exposure. This work presents the development of an immunomagnetic-UHPLC-MS/MS method for the quantitation of unadducted BChE and the long-term CBDP biomarker, oCP-BChE, in human serum. The method has a reportable range from 2.0 ng/ml to 150 ng/ml, which is consistent with the sensitivity of methods used to detect organophosphorus nerve agent protein adducts. The assay demonstrated high intraday and interday accuracy (≥85%) and precision (RSD ≤ 15%) across the calibration range. The method was developed for future analyses of potential human exposure to CBDP. Analysis of human serum inhibited in vitro with CBDP demonstrated that the oCP-BChE adduct was stable for at least 72 h at 4, 22 and 37 °C. Compared to a previously reported assay, this method requires 75% less sample volume, reduces analysis time by a factor of 20 and demonstrates a threefold improvement in sensitivity. Published 2015. This article is a U.S. Government work and is in the public domain in the USA.
authors.affiliation: Oak Ridge Institute for Science and Education, Centers for Disease Control and Prevention, Atlanta, GA, USA.
authors.forename: Darryl
authors.initials: D
authors.lastname: Johnson
authors.affiliation: Division of Laboratory Sciences, National Center for Environmental Health, Centers for Disease Control and Prevention, Atlanta, GA, USA.
authors.forename: Melissa D
authors.initials: MD
authors.lastname: Carter
authors.affiliation: Division of Laboratory Sciences, National Center for Environmental Health, Centers for Disease Control and Prevention, Atlanta, GA, USA.
authors.forename: Brian S
authors.initials: BS
authors.lastname: Crow
authors.affiliation: Oak Ridge Institute for Science and Education, Centers for Disease Control and Prevention, Atlanta, GA, USA.
authors.forename: Samantha L
authors.initials: SL
authors.lastname: Isenberg
authors.affiliation: Oak Ridge Institute for Science and Education, Centers for Disease Control and Prevention, Atlanta, GA, USA.
authors.forename: Leigh Ann
authors.initials: LA
authors.lastname: Graham
authors.affiliation: Oak Ridge Institute for Science and Education, Centers for Disease Control and Prevention, Atlanta, GA, USA.
authors.forename: H Akin
authors.initials: HA
authors.lastname: Erol
authors.affiliation: Oak Ridge Institute for Science and Education, Centers for Disease Control and Prevention, Atlanta, GA, USA.
authors.forename: Caroline M
authors.initials: CM
authors.lastname: Watson
authors.affiliation: Division of Laboratory Sciences, National Center for Environmental Health, Centers for Disease Control and Prevention, Atlanta, GA, USA.
authors.forename: Brooke G
authors.initials: BG
authors.lastname: Pantazides
authors.affiliation: The Netherlands Organization for Applied Scientific Research (TNO), Technical Sciences, CBRN Protection, Rijswijk, The Netherlands.
authors.forename: Marcel J
authors.initials: MJ
authors.lastname: van der Schans
authors.affiliation: The Netherlands Organization for Applied Scientific Research (TNO), Technical Sciences, CBRN Protection, Rijswijk, The Netherlands.
authors.forename: Jan P
authors.initials: JP
authors.lastname: Langenberg
authors.affiliation: The Netherlands Organization for Applied Scientific Research (TNO), Technical Sciences, CBRN Protection, Rijswijk, The Netherlands.
authors.forename: Daan
authors.initials: D
authors.lastname: Noort
authors.affiliation: Division of Laboratory Sciences, National Center for Environmental Health, Centers for Disease Control and Prevention, Atlanta, GA, USA.
authors.forename: Thomas A
authors.initials: TA
authors.lastname: Blake
authors.affiliation: Division of Laboratory Sciences, National Center for Environmental Health, Centers for Disease Control and Prevention, Atlanta, GA, USA.
authors.forename: Jerry D
authors.initials: JD
authors.lastname: Thomas
authors.affiliation: Division of Laboratory Sciences, National Center for Environmental Health, Centers for Disease Control and Prevention, Atlanta, GA, USA.
authors.forename: Rudolph C
authors.initials: RC
authors.lastname: Johnson
mesh_terms: D002091:Butyrylcholinesterase / Q000097:blood* / Q000737:chemistry; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D006801:Humans; D018189:Immunomagnetic Separation / Q000379:methods*; D057230:Limit of Detection; D015203:Reproducibility of Results; D053719:Tandem Mass Spectrometry / Q000379:methods*; D014317:Tritolyl Phosphates / Q000097:blood* / Q000737:chemistry
chemical_list: D014317:Tritolyl Phosphates; D002091:Butyrylcholinesterase; C025541:tri-o-cresyl phosphate
keywords: Jamaica ginger paralysis; butyrylcholinesterase; cresyl saligenin phosphate; organophosphate-induced delayed neuropathy; tri-ortho-cresyl-phosphate