title: Reversed-phase ion-pair chromatography-diode array detection of the bispyridinium compound MB327: plasma analysis of a potential novel antidote for the treatment of organophosphorus poisoning.
abstract: In the case of poisoning by organophosphorus nerve agents or pesticides, there is still a lack of pharmacological treatment of the cholinergic crisis selectively targeting the nicotinic acetylcholine receptor. Recently, the compound MB327 was identified as a potential novel lead structure to close this gap, thus demanding a quantitative assay for initial pharmacokinetic (PK) studies. MB327 is a salt consisting of the dicationic bispyridinium compound (BPC) 1,1´-(propane-1,3-diyl)bis(4-tert-butylpyridinium) and two iodide counter ions. Due to the permanent positive charge of the BPC, an isocratic reversed-phase ion-pair chromatographic separation (RPIPC) was developed using heptanesulfonic acid as ion-pairing reagent and 45% v/v methanol as organic modifier (1 mL/min). Selective UV-detection (230 nm) was done by a diode array detector (DAD) for reliable, rugged, precise (RSD < 7%) and accurate (96-104%) quantitative analysis of 50 μL swine plasma (linear range 1-1000 µg BPC/mL plasma, lower limit of quantification 2 µg/mL). During method validation, diverse parameters essential for the chromatographic process were investigated to generate van´t Hoff, van Deemter and width plots allowing calculation of thermodynamic data like the distribution constant K (5.7 ± 0.3), change in enthalpy, ΔH(0) : -23.66 kJ/mol, and entropy, ΔS(0) : -65 J/(mol*K). In addition, RPIPC-DAD analysis enabled calculation of molar absorptivities of the BPC, ε230 : 17 400 ± 1100 L/(mol*cm), and iodide, ε230 : 9900 ± 400 L/(mol*cm), which determination was hampered by interference with each other in conventional cuvette UV-spectrophotometric measurements. Finally, the RPIPC-DAD procedure was applied to samples from an in vivo study of swine.
authors.affiliation: Bundeswehr Institute of Pharmacology and Toxicology, Munich, Germany.
authors.forename: Harald
authors.initials: H
authors.lastname: John
authors.affiliation: Defence Research and Development Canada-Suffield Research Station, Medicine Hat, Alberta, Canada.
authors.forename: John
authors.initials: J
authors.lastname: Mikler
authors.affiliation: Bundeswehr Institute of Pharmacology and Toxicology, Munich, Germany.
authors.forename: Franz
authors.initials: F
authors.lastname: Worek
authors.affiliation: Bundeswehr Institute of Pharmacology and Toxicology, Munich, Germany.
authors.forename: Horst
authors.initials: H
authors.lastname: Thiermann
mesh_terms: D000818:Animals; D000931:Antidotes / Q000032:analysis* / Q000627:therapeutic use; D001342:Autonomic Nervous System Diseases / Q000139:chemically induced / Q000188:drug therapy; D002619:Chemical Warfare Agents / Q000506:poisoning; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D056148:Chromatography, Reverse-Phase; D008297:Male; D062025:Organophosphate Poisoning / Q000188:drug therapy*; D009946:Organothiophosphorus Compounds / Q000506:poisoning; D011726:Pyridinium Compounds / Q000097:blood* / Q000627:therapeutic use; D011978:Receptors, Nicotinic / Q000187:drug effects; D015203:Reproducibility of Results; D013056:Spectrophotometry, Ultraviolet; D013552:Swine
chemical_list: D000931:Antidotes; D002619:Chemical Warfare Agents; D009946:Organothiophosphorus Compounds; D011726:Pyridinium Compounds; D011978:Receptors, Nicotinic; C571852:MB327; C009680:VX-agent
keywords: cholinergic crisis; ion-pair chromatography; molar absorptivities; nerve agent poisoning; van Deemter plot; van´t Hoff plot