Simultaneous determination of 3-mercaptopyruvate and cobinamide in plasma by liquid chromatography-tandem mass spectrometry.

The current suite of Food and Drug Administration (FDA) approved antidotes (i.e., sodium nitrite, sodium thiosulfate, and hydroxocobalamin) are effective for treating cyanide poisoning, but individually, each antidote has major limitations (e.g., large effective dosage or delayed onset of action). To mitigate these limitations, next-generation cyanide antidotes are being investigated, including 3-mercaptopyruvate (3-MP) and cobinamide (Cbi). Analytical methods capable of detecting these therapeutics individually and simultaneously (for combination therapy) are essential for the development of 3-MP and Cbi as potential cyanide antidotes. Therefore, a liquid chromatography-tandem mass-spectrometry method for the simultaneous analysis of 3-MP and Cbi was developed. Sample preparation of 3-MP consisted of spiking plasma with an internal standard ((13)C3-3-MP), precipitation of plasma proteins, and derivatizing 3-MP with monobromobimane to produce 3-mercaptopyruvate-bimane. Preparation of Cbi involved denaturing plasma proteins with simultaneous addition of excess cyanide to convert each Cbi species to dicyanocobinamide (Cbi(CN)2). The limits of detection for 3-MP and Cbi were 0.5µM and 0.2µM, respectively. The linear ranges were 2-500µM for 3-MP and 0.5-50µM for Cbi. The accuracy and precision for 3-MP were 100±9% and <8.3% relative standard deviation (RSD), respectively. For Cbi(CN)2, the accuracy was 100±13% and the precision was <9.5% RSD. The method presented here was used to determine 3-MP and Cbi from treated animals and may ultimately facilitate FDA approval of these antidotes for treatment of cyanide poisoning.

Synthesis and characterization of a new binucleating tetraphosphine ligand based on 1,2-phenylene chelates and the structures of dinickel tetrachloride complexes of the ligand.

A new binucleating tetraphosphine ligand, rac- and meso-(Et2P-1,2-C6H4)P(Ph)CH2(Ph)P(1,2-C6H4PEt2) (et,ph-P4-Ph), has been synthesized. Separation and purification of the ligand diastereomers have been accomplished via column chromatography. Ni2Cl4(et,ph-P4-Ph) complexes of both diastereomers have been prepared in high yield and crystallographically characterized.

Determination of 3-mercaptopyruvate in rabbit plasma by high performance liquid chromatography tandem mass spectrometry.

Accidental or intentional cyanide poisoning is a serious health risk. The current suite of FDA approved antidotes, including hydroxocobalamin, sodium nitrite, and sodium thiosulfate is effective, but each antidote has specific major limitations, such as large effective dosage or delayed onset of action. Therefore, next generation cyanide antidotes are being investigated to mitigate these limitations. One such antidote, 3-mercaptopyruvate (3-MP), detoxifies cyanide by acting as a sulfur donor to convert cyanide into thiocyanate, a relatively nontoxic cyanide metabolite. An analytical method capable of detecting 3-MP in biological fluids is essential for the development of 3-MP as a potential antidote. Therefore, a high performance liquid chromatography tandem mass spectrometry (HPLC-MS-MS) method was established to analyze 3-MP from rabbit plasma. Sample preparation consisted of spiking the plasma with an internal standard ((13)C3-3-MP), precipitation of plasma proteins, and reaction with monobromobimane to inhibit the characteristic dimerization of 3-MP. The method produced a limit of detection of 0.1µM, a linear dynamic range of 0.5-100µM, along with excellent linearity (R(2)≥0.999), accuracy (±9% of the nominal concentration) and precision (<7% relative standard deviation). The optimized HPLC-MS-MS method was capable of detecting 3-MP in rabbits that were administered sulfanegen, a prodrug of 3-MP, following cyanide exposure. Considering the excellent performance of this method, it will be utilized for further investigations of this promising cyanide antidote.

Cyanide antidotes for mass casualties: water-soluble salts of the dithiane (sulfanegen) from 3-mercaptopyruvate for intramuscular administration.

Current cyanide antidotes are administered by IV infusion, which is suboptimal for mass casualties. Therefore, in a cyanide disaster, intramuscular (IM) injectable antidotes would be more appropriate. We report the discovery of the highly water-soluble sulfanegen triethanolamine as a promising lead for development as an IM injectable cyanide antidote.