Development of an LC–Tandem Mass Spectrometry Method for the Quantitative Analysis of Hercynine in Human Whole Blood

Salvatore Sotgia, Rhys B. Murphy, Angelo Zinellu, David Elliot, Panagiotis Paliogiannis, Gerard Aime Pinna, Ciriaco Carru, Arduino A. Mangoni

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Given that the peculiar redox behavior of ergothioneine involves a rapid regeneration process, the measurement of its precursor and redox metabolite hercynine could be particularly useful in assessing its role in oxidative stress or other biological processes. Thus, a LC-MS/MS method for the determination of hercynine concentrations in whole blood was developed. After lysis of red blood cells by cold water, samples were filtered on micro concentrators at a controlled temperature of 4 C. The clear filtered fluid was then treated with diethylpyrocarbonate to derivatize hercynine for the analysis by LC-MS/MS. The derivatized analyte was isocratically separated as a carbethoxy derivative on a C18 column with a mobile phase of an aqueous 0.1% v/v formic acid and acetonitrile (95:5). Effluents were monitored by MRM transitions at m/z 270.28→95 and 273.21→95 for hercynine and its deuterated counterpart, respectively. No cross-talk between MRM transitions was observed and a good linearity was found within a range of 35–1120 nmol/L. The LOD and LOQ were, respectively, 10.30 and 31.21 nmol/L with an intraday and intermediate precision below 7%. The average hercynine concentration in whole blood from 30 healthy male volunteers (aged 77 ± 12 years) was 178.5 ± 118.1 nmol/L. Overall, the method is easy to perform, allowing a rapid and accurate assessment of whole blood concentrations of hercynine.

Original languageEnglish
Article number3326
Number of pages10
Issue number12
Publication statusPublished - 14 Dec 2018


  • Aminothione
  • Antioxidant
  • Dismutatiion
  • LC-tandem mass spectrometry
  • Zwitterion


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