Enzyme self-inactivation is a main limitation of the preparation of eicosanoids - Enzymatic synthesis of thromboxane and 12(S)-hydroxytetraenoic acid

Synthesis of prostanoids is accompanied by various processes reducing the p roduct yield. These processes are also known to affect syntheses of thrombo xane (TX) and 12(S)-hydroxy-5(Z),8(Z),10(E),14(Z)-eicosatetraenoic acid (12 -HETE). Partially purified preparations of TX synthase and prostaglandin (P G) synthase were used to optimize TX synthesis with respect to concentratio ns of the enzymes and eicosapolyenoic acid (EPA). Conditions for the maximu m product yield and the minimum consumption of enzymes were determined. Con sumption of the TX synthase was large owing to its inactivation during the reaction and the nonenzymatic destruction of the intermediate product PG-en doperoxide. Separate addition of PG and TX synthases increased the product yield by preventing EPA sorption on ballast proteins. Microsomal 12-lipoxyg enase (12-LO) was also shown to be inactivated during the reaction, and thi s process was the major limitation of 12-HETE synthesis. Lipoxygenase react ion in the presence of some reducing agents led to a considerable increase of the 12-HETE yield, supposedly by preventing further oxidation of the 12- LO reaction product 12-hydroperoxy derivative of eicosatetraenoic acid. The possibility of using human blood platelet microsomes for preparation of so me derivatives of EPAs is discussed.