Archives of Toxicology

, Volume 78, Issue 9, pp 508–524 | Cite as

Retrospective detection of exposure to nerve agents: analysis of phosphofluoridates originating from fluoride-induced reactivation of phosphylated BuChE

  • Marcel J. van der Schans
  • Martine Polhuijs
  • Corry van Dijk
  • Carla E. A. M. Degenhardt
  • Kees Pleijsier
  • Jan P. Langenberg
  • Hendrik P. Benschop
Organ Toxicity and Mechanisms

Abstract

The utility was explored of a new approach to detect retrospectively exposure to nerve agents by means of conversion of the inhibitor moiety bound to the active site of the enzyme BuChE in plasma with fluoride ions into a phosphofluoridate which is subsequently analyzed by means of gas chromatography (GC). This quantifies ≥0.01% inhibition of BuChE and identifies the structure of the inhibitor except for the original leaving group. A three-tiered approach was followed involving the five classical nerve agents GA, GB, GF, GD, and VX, as well as the active metabolite of parathion, i.e., paraoxon: in vivo experiments in rhesus monkeys after iv administration of a sign-free dose of agent and concomitant in vitro experiments in plasma of rhesus monkeys and humans should allow an assessment of in vivo retrospectivity in humans. A systematic investigation was performed in order to find a single set of reaction conditions which yields a maximum amount of phosphofluoridate for all nerve agents. Fluoride-induced reactivation at 25°C at a final concentration of 250 mM KF during 15 min in a pH-range between 4 and 6 appears to be effective. The in vitro decrease with time in reactivatibility of inhibited BuChE in plasma from humans and rhesus monkeys was largely due to aging of the phosphyl moiety, except for VX where spontaneous reactivation was a major cause. The decrease followed first-order except for a biphasic course in the case of GF in human and rhesus monkey plasma as well as of GD in rhesus plasma. In vitro half-lifes in human plasma ranged between ca. 14 h for GB and ca. 63 h for GA. A comparison of the in vivo data from rhesus monkeys and the in vitro data is complicated by the observation that the in vivo decrease with time of fluoride-reactivated phosphofluoridate is biphasic for all nerve agents. The terminal in vivo phase pertains to a small fraction of the amount of initially regenerated phosphofluoridate but is responsible for a considerable degree of retrospectivity, ranging between 14 and 56 days for GF and GB, respectively. The new procedure can be used in a variety of practical applications, e.g., (i) biomonitoring in health surveillance at exposure levels that are several orders of magnitude lower than presently possible; (ii) diagnosis in case of alleged exposure to nerve agents in time of war or after terrorist attacks; (iii) in forensic cases against suspected terrorists that have handled organophosphate anticholinesterases; and (iv) in research applications such as investigations on lowest observable effect levels of exposure to nerve agents.

Keywords

Biomarker Biomonitoring Butyryl cholinesterase Fluoride ions Low-level exposure Nerve agents Organophosphates Reactivation Retrospective detection 

Abbreviations

AChE

Acetylcholinesterase

BuChE

Butyrylcholinesterase

BuSChI

Butyrylthiocholine iodide

DEFP

O,O-diethyl phosphorofluoridate

DFP

O,O-diisopropyl phosphorofluoridate

CaE

Carboxylesterase

CI

Chemical ionisation

CIS

Cooled injection system

CW

Chemical warfare

ES

O-ethyl methylphosphonofluoridate, ethylsarin

FT

O-ethyl dimethylphosphoramidofluoridate, fluorotabun

GA

O-ethyl dimethylphosphoramidocyanidate, tabun

GB

O-isopropyl methylphosphonofluoridate, sarin

GC

Gas chromatography

GD

O-pinacolyl methylphosphonofluoridate, soman

GF

O-cyclohexyl methylphosphonofluoridate, cyclohexyl sarin

LC

Liquid chromatography

MS

Mass spectrometry

MSD

Mass selective detection

MUSIC

Multiple switchable intelligent controller

NP

Detection nitrogen phosphorus selective detection

PFPD

Pulsed flame photometric detection

TCT

Thermodesorption cold trap

TDSA

Thermodesorption autosampler

VX

O-ethyl S-2-diisopropylaminoethyl methylphosphonothioate

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Copyright information

© Springer-Verlag 2004

Authors and Affiliations

  • Marcel J. van der Schans
    • 1
  • Martine Polhuijs
    • 1
  • Corry van Dijk
    • 1
  • Carla E. A. M. Degenhardt
    • 1
  • Kees Pleijsier
    • 1
  • Jan P. Langenberg
    • 1
  • Hendrik P. Benschop
    • 1
  1. 1.Department of Medical CountermeasuresTNO Prins Maurits LaboratoryRijswijkThe Netherlands

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