prostaglandin-a1 and lauric-acid

We have examined in detail the substrate specificity of a rabbit kidney fatty acid omega-hydroxylase, designated cytochrome P-450ka2 (CYP4A7). The hydroxylation products were identified as omega- and (omega - 1)-hydroxy fatty acids mainly using gas chromatography-electron impact mass spectrometry. [1] Straight-chain saturated fatty acids ranging from 10 to 19 carbons were effectively hydroxylated at the omega- and (omega - 1)-position. The ratios of omega- to (omega - 1)-hydroxylation activity decreased with increasing the carbon chain length of fatty acids. [2] Both isomyristate and anteisomyristate, and isopalmitate were hydroxylated several fold more rapidly than myristate and palmitate, respectively, with iso-branched chain fatty acids being hydroxylated at the omega-position solely. [3] Both palmitoleate and palmitoelaidate, and both oleate and elaidate were hydroxylated much more rapidly than palmitate and stearate, respectively. [4] Linoleate, gamma-linolenate, and arachidonate were also excellent substrates for this enzyme. [5] Prostaglandin (PG) A1 and PGA2 were efficiently hydroxylated at the omega-position solely, with PGE1 and PGE2 being much less active. [6] Arachidonic acid not only showed a Km value significantly lower than those for lauric acid, gamma-linolenic acid and PGA1, but also it is a potent competitor for lauric acid and PGA1, showing a very high affinity for the enzyme. It is possible that arachidonic acid is the physiological substrate for kidney P-450ka2.

Topics: Animals; Arachidonic Acid; Catalysis; Cytochrome P-450 CYP4A; Cytochrome P-450 Enzyme System; Cytochromes b5; Fatty Acids; Hydroxylation; Kidney Cortex; Kinetics; Lauric Acids; Microsomes; Mixed Function Oxygenases; Phosphatidylcholines; Prostaglandins A; Rabbits; Substrate Specificity

Terminal acetylenic fatty acid mechanism-based inhibitors (Ortiz de Montellano, P. R., and Reich, N. O. (1984) J. Biol. Chem. 259, 4136-4141) were used as probes in determining the substrate specificity of rabbit lung cytochrome P-450 isozymes of pregnant animals in both microsomes and reconstituted systems. Lung microsomal and reconstituted P-450 form 5-catalyzed lauric acid omega- and (omega-1)-hydroxylase activities were inhibited by a 12-carbon terminal acetylenic fatty acid, 11-dodecynoic acid (11-DDYA), and an 18-carbon terminal acetylenic fatty acid, 17-octadecynoic acid (17-ODYA). Rabbit lung microsomal lauric acid omega-hydroxylase activity was more sensitive to inhibition by 11-DDYA than was (omega-1)-hydroxylase activity. In reconstituted systems containing purified P-450 form 5, both omega- and (omega-1)-hydroxylation of lauric acid were inhibited in parallel when either 11-DDYA or 17-ODYA was used. These data suggest the presence of at least two P-450 isozymes in rabbit lung microsomes capable of lauric acid omega-hydroxylation. This is the first report indicating the multiplicity of lauric acid hydroxylases in lung microsomes. Lung microsomal prostaglandin omega-hydroxylation, mediated by the pregnancy-inducible P-450PG-omega (Williams, D. E., Hale, S. E., Okita, R. T., and Masters, B. S. S. (1984) J. Biol. Chem. 259, 14600-14608) was subject to inhibition by 17-ODYA only, whereas 11-DDYA acid was not an effective inhibitor of this hydroxylase. We have recently developed a new terminal acetylenic fatty acid, 12-hydroxy-16-heptadecynoic acid (12-HHDYA), that contains a hydroxyl group at the omega-6 position. We show that 12-HHDYA possesses a high degree of selectivity for the inactivation of rabbit lung microsomal prostaglandin omega-hydroxylase activity which cannot be obtained with the long chain acetylenic inhibitor, 17-ODYA. In addition, 12-HHDYA has no effect on lauric acid omega- or omega-1-hydroxylation or on benzphetamine N-demethylation. The development of this new terminal acetylenic fatty acid inhibitor provides us with a useful tool with which to study the physiological role of prostaglandin omega-hydroxylation in the rabbit lung during pregnancy.

Topics: Animals; Cytochrome P-450 CYP4A; Cytochrome P-450 Enzyme System; Dinoprostone; Fatty Acids, Unsaturated; Isoenzymes; Kinetics; Lauric Acids; Lung; Microsomes; Mixed Function Oxygenases; Prostaglandins A; Rabbits

The effect of Lipo PGE1 (prostaglandin E1 incorporated in lipid microspheres) on laurate-induced peripheral arterial occlusive disease in rats was evaluated and compared with that of other related compounds. Lipo PGE1 showed a potent inhibitory effect on the progression of lesions in this model, but Lipo PGA1 had no such effect. Moreover, the effect of Lipo PGE1 was more potent than that of PGE1 X CD (cyclodextrin clathrated PGE1) and PGE1 on this model. These results suggest that incorporation of PGE1 into soybean oil microspheres might exert a more potent effect by facilitating drug delivery to the lesions in this model. It would appear that Lipo PGE1 is suitable for clinical use in peripheral vascular disorders.

Topics: Alprostadil; Animals; Arterial Occlusive Diseases; Dosage Forms; Lauric Acids; Male; Microspheres; Prostaglandins A; Rats; Rats, Inbred Strains