TY - JOUR
T1 - Liquid chromatography-tandem mass spectrometry for simultaneous measurement of thromboxane B2 and 12(S)-hydroxyeicosatetraenoic acid in serum
AU - Squellerio, Isabella
AU - Porro, Benedetta
AU - Songia, Paola
AU - Veglia, Fabrizio
AU - Caruso, Donatella
AU - Tremoli, Elena
AU - Cavalca, Viviana
PY - 2014/8/5
Y1 - 2014/8/5
N2 - Arachidonic acid (AA) is metabolized in human platelets by two main pathways: via cyclooxygenase (COX-1) to prostaglandins and thromboxane (TX)A2 and via 12-lipoxygenase (12-LOX) to 12(S)-hydroxyeicosatetraenoic acid (12(S)-HETE). While COX products are known to regulate platelet reactivity, the role of 12-LOX metabolites is still controversial. To better understand the platelet enzymatic pathways, we developed a simple and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay for the simultaneous measurement of both platelet metabolites in human serum. After the addition of deuterated d4-TXB2 and d8-12(S)-HETE as internal standards and the solid-phase extraction of serum samples, analytes were resolved using reversed-phase C18 column and quantified using negative ion electrospray ionization-tandem mass spectrometry. Intra and interassay imprecisions were less than 10% for both analytes. The lower limits of quantification were 0.244ng/ml and 0.976ng/ml for TXB2 and 12(S)-HETE, respectively. This method was applied to measure platelet metabolites in healthy subjects (n=35). LC-MS/MS allows rapid, simultaneous, sensitive and accurate quantification of both platelet AA products in human serum with a small sample volume required and a minimal sample preparation.
AB - Arachidonic acid (AA) is metabolized in human platelets by two main pathways: via cyclooxygenase (COX-1) to prostaglandins and thromboxane (TX)A2 and via 12-lipoxygenase (12-LOX) to 12(S)-hydroxyeicosatetraenoic acid (12(S)-HETE). While COX products are known to regulate platelet reactivity, the role of 12-LOX metabolites is still controversial. To better understand the platelet enzymatic pathways, we developed a simple and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay for the simultaneous measurement of both platelet metabolites in human serum. After the addition of deuterated d4-TXB2 and d8-12(S)-HETE as internal standards and the solid-phase extraction of serum samples, analytes were resolved using reversed-phase C18 column and quantified using negative ion electrospray ionization-tandem mass spectrometry. Intra and interassay imprecisions were less than 10% for both analytes. The lower limits of quantification were 0.244ng/ml and 0.976ng/ml for TXB2 and 12(S)-HETE, respectively. This method was applied to measure platelet metabolites in healthy subjects (n=35). LC-MS/MS allows rapid, simultaneous, sensitive and accurate quantification of both platelet AA products in human serum with a small sample volume required and a minimal sample preparation.
KW - 12(S)-hydroxyeicosatetraenoic acid
KW - Mass spectrometry
KW - Platelet
KW - Serum
UR - http://www.scopus.com/inward/record.url?scp=84899680352&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84899680352&partnerID=8YFLogxK
U2 - 10.1016/j.jpba.2014.04.004
DO - 10.1016/j.jpba.2014.04.004
M3 - Article
C2 - 24786190
AN - SCOPUS:84899680352
VL - 96
SP - 256
EP - 262
JO - Journal of Pharmaceutical and Biomedical Analysis
JF - Journal of Pharmaceutical and Biomedical Analysis
SN - 0731-7085
ER -