MICRODETERMINATION OF 11-DEHYDROTHROMBOXANE B-3 IN HUMAN URINE BY GAS-CHROMATOGRAPHY SELECTED-ION MONITORING USING [O-18(2)] ANALOG AS AN INTERNAL STANDARD

The microdetermination of 11-dehydrothromboxane B-3 (11-dehydro-TXB(3) ) in human urine is described, We prepared [O-18(2)]11-dehydrothrombox ane B-3 for use as an internal standard. Samples containing an [O-18(2 )] analogue were extracted chromatographically using Sep Pak tC18 and a silica gel column. Conversion of the extracted 11-dehydro-TXB(3) to 1-methyl er-11-n-propylamide-9,12,15-dimethylisopropylsilyl ether deri vative was followed by gas chromatography/selected ion monitoring (GC/ SIM). Interfering substances from the urine matrix were eliminated dur ing GC/SIM analysis using an MP-65HT column. Human urine was monitored using the ions of mit 696.4511 (11-dehydro-TXB(3)), m/z 700.4597 ([O- 18(2)]11-dehydro-TXB(3)), m/z 698.4668 (11-dehydro-TXB(2)) and m/z 702 .4918 ([H-2(4)]11-dehydro-TXB(2)). 11-Dehydro-TXB(3) and [O-18(2)]11-d ehydro-TXB(3) appeared at the same retention time and were eluted 15 s after 11-dehydro-TXB(3) on this capillary column. This difference in retention time was due to the double bond in the omega 3 position in 1 1-dehydro-TXB(3). A good linear response over the range of 10 pg to 10 ng/tube was demonstrated. We detected 11-dehydro-TXB(3) in the range from 1.29 to 7.64 pg/mg creatinine in the human urine. In the healthy volunteers, the 11-dehydro-TXB(3) level was less than 1% of that of th e 11-dehydro-TXB(2), but 11-dehydro-TXB(3) was increased by dietary su pplementation of eicosapentaenoic acid. This method can be applied to the determination of 11-dehydro-TXB(3), in human urine.