Arachidonic acid is responsible for the smooth muscle-contracting activity of G-acid.

Abstract

G-acid, a smooth-muscle-contracting substance isolated from human blood plasma, was previously identified as 3-octadecenoic acid. Synthetic cis-and trans-3-octadecenoic acid, however, are biologically inactive. C-acid actually consists of a mixture of fatty acids, characteristic of animal tissues. The biological activity is caused by arachidonic acid, which contracts smooth muscle upon in situ conversion to prostaglandins.In 1956, Gabr reported the isolation of a substance from human blood plasma which slowly contracted isolated guinea pig jejunum or ileum and was thought to cause certain adverse plasma transfusion reactions (1, 2). He named this substance, associated with the globulin fraction, G-acid, and deduced that it was a fatty acid containing 18 carbons and one double bond. G-acid was tentatively characterized as 3-octadecenoic acid on the basis of its oxidation with KMnO4, which produced malonic acid. Subsequently, various authors have questioned the validity of this structural assignment (3, 4), but no re-examination has been reported.We have synthesized cis-and trans-3-octadecenoic acid and established that both compounds are biologically inactive. Our study reveals that Gabr's G-acid consists of a mixture of fatty acids quite characteristic of those commonly found in animal tissues. All of the biological activity is due to the arachidonic acid (all cis 5,8,11,14-eicosatetraenoic acid) present in this mixture.MATERIALS AND METHODS Bioassay. Fatty acids, dispersed in 1 ml of Tyrode's solution, were added to isolated guinea pig jejunum (1) or ileum (2) bathed in 9 ml of Tyrode's solution. Atropine-free Tyrode's solution was used unless otherwise stated. When included, the concentration of atropine sulfate was 35 ng/ml. Ileum (2), which is more sensitive than jejunum, was used for routine assays, but all significant results were also confirmed with jejunum. Contractions were recorded with a low-friction writing lever on carbon-coated paper. Contractions generally began 15-30 sec after arachidonic acid or G-acid samples were applied and reached a maximum after about 2 min. Approximately equal contractions were produced by (per ml) 40 ng of serotonin, 150 ng of arachidonic acid, 2 ng of prostaglandin E2 (PGE2), and 25 ng of prostaglandin F2a (PGF2,,).cis-and trans-3-Octadecenoic Acid. A modification of the procedure of Rando and Doering (5) was used. Methyl 2-octadecenoate (6) (105 mg, -0.34 mmol), dissolved in 1 ml of ethyl acetate and 19 ml of methanol, was irradiated in a nitrogen atmosphere at 100C for 3 hr with an unfiltered mercury arc lamp. Preparative thin-layer chromatography (TLC) on silica gel (hexane/ether, 85:15 vol/vol) afforded a mixture of methyl cis-and trans-3-octadecenoate (RF = 0.45, 24 mg). Subsequent chromatography of these esters on AgNO3 (20%)-impregnated silica gel (toluene, -250C) (7) separated the cis (4 mg, 4% yield) from the trans (12 mg, 12% yield) isomer. The esters were hydrolyzed for 10 min with 0.5 M NaOH in ethanol/water, 9:1 (vol/vol). After acidification to pH 2...