Quantitation of Ortho-cresyl Phosphate Adducts to Butyrylcholinesterase in Human Serum by Immunomagnetic-UHPLC-MS/MS

Authors

• Darryl Johnson, United States Centers for Disease Control and Prevention
• Melissa D. Carter, United States Centers for Disease Control and PreventionFollow
• Brian S. Crow, United States Centers for Disease Control and Prevention
• Samantha L. Isenberg, United States Centers for Disease Control and Prevention
• Leigh Ann Graham, United States Centers for Disease Control and Prevention
• H. Akin Erol, United States Centers for Disease Control and Prevention
• Caroline M. Watson, United States Centers for Disease Control and Prevention
• Brooke G. Pantazides, United States Centers for Disease Control and Prevention
• Marcel J. van der Schans, Netherlands Organization for Applied Scientific Research
• Jan P. Langenberg, Netherlands Organization for Applied Scientific Research
• Daan Noort, Netherlands Organization for Applied Scientific Research
• Thomas A. Blake, United States Centers for Disease Control and Prevention
• Jerry D. Thomas, United States Centers for Disease Control and Prevention
• Rudolph C. Johnson, United States Centers for Disease Control and Prevention

Date of this Version

1-2015

Citation

Journal of Mass Spectrometry (2015) 50: 683–692

doi: 10.1002/jms.3576

Comments

United States government work

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 fromthe 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 2.0ng/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 72h 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.