Mikael Lindström1, Miia Valkonen2, Niina Tohmola3, Risto Renkonen4, Tomas Strandin5, Antti Vaheri5, Outi Itkonen3. 1. HUSLAB, University of Helsinki and Helsinki University Hospital, Helsinki, Finland. Electronic address: mikael.lindstrom@hus.fi. 2. Division of Intensive Care Medicine, Department of Anesthesiology, Intensive Care and Pain Medicine, University of Helsinki and Helsinki University Hospital, Helsinki, Finland. 3. HUSLAB, University of Helsinki and Helsinki University Hospital, Helsinki, Finland. 4. HUSLAB, University of Helsinki and Helsinki University Hospital, Helsinki, Finland; Faculty of Medicine, University of Helsinki, Finland. 5. Faculty of Medicine, University of Helsinki, Finland.
Abstract
BACKGROUND: Bradykinin is an important mediator of inflammation and vascular permeability and could have an important role in the development of septic shock. Measurement of bradykinin by immunological methods may suffer from interference and lack of specificity. We developed and validated a liquid chromatography mass spectrometry assay (LC-MS/MS) for plasma bradykinin. METHODS: We used plasma samples from healthy volunteers (n = 19) and patients with septic shock (n = 47). Stable isotope bradykinin internal standard was added to samples before solid-phase extraction and quantification by LC-MS/MS. Stability of bradykinin was studied for 12 months. RESULTS: Our assay has good sensitivity (0.1 nmol/l) and a wide linear range (0.1-1000 nmol/l). Bradykinin added to plasma was stable for 12 months at -20 °C when a mixture of protease inhibitors was added at sampling but degraded during repeated freezing and thawing. Bradykinin concentration in plasma from septic shock patients (<0.1-0.6 nmol/l) did not change significantly during shock and recovery but differed slightly from that in healthy individuals (0.5-1.1 nmol/l). CONCLUSIONS: Our bradykinin assay was successfully used to determine bradykinin concentrations in plasma samples. Intensive care unit patients with septic shock had low concentrations of plasma bradykinin during both shock and recovery phases.
BACKGROUND:Bradykinin is an important mediator of inflammation and vascular permeability and could have an important role in the development of septic shock. Measurement of bradykinin by immunological methods may suffer from interference and lack of specificity. We developed and validated a liquid chromatography mass spectrometry assay (LC-MS/MS) for plasma bradykinin. METHODS: We used plasma samples from healthy volunteers (n = 19) and patients with septic shock (n = 47). Stable isotope bradykinin internal standard was added to samples before solid-phase extraction and quantification by LC-MS/MS. Stability of bradykinin was studied for 12 months. RESULTS: Our assay has good sensitivity (0.1 nmol/l) and a wide linear range (0.1-1000 nmol/l). Bradykinin added to plasma was stable for 12 months at -20 °C when a mixture of protease inhibitors was added at sampling but degraded during repeated freezing and thawing. Bradykinin concentration in plasma from septic shockpatients (<0.1-0.6 nmol/l) did not change significantly during shock and recovery but differed slightly from that in healthy individuals (0.5-1.1 nmol/l). CONCLUSIONS: Our bradykinin assay was successfully used to determine bradykinin concentrations in plasma samples. Intensive care unit patients with septic shock had low concentrations of plasma bradykinin during both shock and recovery phases.