簡便な11-dehydro-thromboxane B2の抽出法の確立とRIAによる測定

Quantitative analysis of thromboxane (TX) levels in vivo is a useful approach to clerify clinical feature of cardiovascular disease. TXA2 is metabolized to TXB2 at nonenzymatic. Consequently plasma TXB2 level is fluctuated by TXA2 which was produced from platelets during blood collection. 11-dehydro-thromboxane B2 (11 DT) is a major metabolite of TXB2 and not producedby the stimulation of platelets during blood collection.We studied a simple and rapid extraction method for 11 DT. 11 DT was extracted from sample by the addition of octadecylsilyl silica powder (80 mg/ml) suspended in 40% aqueous EtOH followed by the fractionation using silica open minicolumn BOND ELUT Si (Analytichem International, Inc., Harbor City, USA) solutions in three different composition of acetonitrile, chloroform and acetic acid were used as eluents. These fractions were quantitated by radioimmunoassay (NEN, Boston USA) .11 DT was recovered 76.1±2.3% (n=10) as closed form using the both of open and closed form of [3H] -11 DT as tracer. Concentrated plasma 11 DT obtained by this method in normal subject was less than 5.2 pg/ml (n=130, male; n=64 age 1948, female; n=66 age 18-48) . There was no sexual difference in plasma 11 DT level by this method. Plasma 11 DT levelwas not so high as that of TXB2 which was directly influenced by the platelet release during blood collection. Serum 11 DT level was 2, 400±500 pg/ml (n=10) which is higher than that in plasma (7.1±1.9 pg/ml) .In conclusion, the technical advantages of this method can be summarized into two major points, simplicity and requiring small volume of sample.