[Marine fungus Stilbella aciculosa as a potential producer of prostaglandins].

The amount and composition of fatty acids in the fungus Stilbella aciculosa associated with the marine macroorganism Apostichopus japonica (trepang) were determined by gas-liquid chromatography and gas chromatography-mass spectrometry. In the culture liquid of S. aciculosa, prostaglandins (PG) of groups E and F were revealed by UV spectroscopy. This finding was confirmed by the presence of direct precursors of PG, polyunsaturated eicosapentaenoic and docosahexaenoic acids, in the culture liquid. The biomass of this fungus contained PG of group B.

Prostaglandin biosynthesis by midgut tissue isolated from the tobacco hornworm, Manduca sexta.

We describe prostaglandin (PG) biosynthesis by isolated midgut preparations from tobacco hornworms, Manduca sexta. Microsomal-enriched midgut preparations yielded four PGs, PGA/B(2), PGD(2), PGE(2) and PGF(2alpha), all of which were confirmed by analysis on gas chromatography--mass spectrometry (GC--MS). PGA and PGB are double bond isomers which do not resolve on TLC but do resolve by GC; for convenience, we use the single term PGA(2) for this product. PGA(2) was the major product under most conditions. The midgut preparations were sensitive to reaction conditions, including radioactive substrate, protein concentration (optimal at 1mg/reaction), reaction time (optimal at 0.5 min), temperature (optimal at 22 degrees C), buffer pH (highest at pH 6), and the presence of a co-factor cocktail composed of reduced glutathione, hydroquinine and hemoglobin. In vitro PG biosynthesis was inhibited by two cyclooxygenase inhibitors, indomethacin and naproxen. Subcellular localization of PG biosynthetic activity in midgut preparations, determined by ultracentrifugation, revealed the presence of PG biosynthetic activity in the cytosolic and microsomal fractions, although most activity was found in the cytosolic fractions. This is similar to other invertebrates, and different from mammalian preparations, in which the activity is exclusively associated with the microsomal fractions. Midgut preparations from M. sexta pupae, adult cockroach, Periplaneta americana, and corn ear worms, Helicoverpa zea, also produced the same four major PG products. We infer that insect midguts are competent to biosynthesize PGs, and speculate they exert important, albeit unrevealed, actions in midgut physiology.

Biosynthesis of prostaglandins from 17(18)epoxy-eicosatetraenoic acid, a cytochrome P-450 metabolite of eicosapentaenoic acid.

Eicosapentaenoic acid (20:5(n - 3)) is oxygenated to 17S(18R)epoxyeicosatetraenoic acid (EpETE) by microsomes of monkey seminal vesicles, which also are rich in prostaglandin (PG) H synthase. The metabolism of racemic [14C]17(18)EpETE by PGH synthase of sheep vesicular glands was investigated in the present report. The two main metabolites were identified by GC-MS as 17(18)epoxyprostagland E2 (17(18)EpPGE2) and 17(18)EpPGF2 alpha. The structures were confirmed by chemical synthesis of these prostaglandins from PGE3. 17(18)EpPGE1 was synthesized from 17,18-dehydro-PGE1 by the same method. Alkali treatment of 17(18)EpPGE2 yielded 17(18)EpPGB2, which could be resolved by RP-HPLC into the 17R(18S) and 17S(18R) stereoisomers. The 17S(18R) stereoisomer was identified by co-chromatography with [14C]17S(18R)EpPGB2, which was formed by PGH synthase from biosynthetic [14C]17S(18R)EpETE. The 17(18)epoxyprostaglandins were found to be relatively unstable during acidic extractive isolation. 17(18)EpPGE1 and 17(18)EpPGE2 could not be detected in seminal vesicles of the cynomolgus monkey in significant amounts relative to 19-hydroxy-PGE1. Nevertheless, biosynthesis of 17(18)epoxyprostaglandins should be considered when the biological effects of 17S(18R)EpETE are investigated.

Metabolism of 5(6)-expoxyeicosatrienoic acid by ram seminal vesicles. Formation of novel prostaglandin E1 metabolites.

5(6)-Epoxy-8,11,14-eicosatrienoic acid was incubated with microsomes or ram seminal vesicles in the presence of glutathione (1 mM) for 2 min at 37 degrees C. Following extractive isolation on octadecasilane silica, the products were purified on straight-phase HPLC and separated into three major polar metabolites, which all showed maximal ultraviolet absorbance at 278 nm after treatment with alkali. The least-polar of the three metabolites was identified by capillary column gas chromatography-mass spectrometry as 5(6)- epoxyprostaglandin E1 and the structure was confirmed by comparison with authentic material. The most-polar metabolite was identified as 5,6- dihydroxyprostaglandin E1, while the metabolite of medium polarity was identified as its delta 5-lactone. When glutathione was omitted, 5-hydroxyprostaglandin I 1 alpha and 5-hydroxyprostaglandin I 1 beta were previously identified as the two major metabolites. These results indicate that the postulated epoxyprostaglandin endoperoxide intermediates, 5(6)- epoxyprostaglandin G1 and 5(6)- epoxyprostaglandin H1, might be substrates for the endoperoxide E isomerase enzyme, since this enzyme requires glutathione as a cofactor.

[Demonstration of the biosynthesis of prostaglandins from arachidonic acid by rat adrenal cortex cells in culture]. / Mise en évidence de la biosynthèse de prostaglandines à partir d'acide arachidonique par les cellules corticosurrénaliennes de rat en culture.

The biosynthesis of prostaglandins of the E pathway was shown in new-born rat adrenocortical cells in long-term primary culture, by direct incubation of labelled arachidonic acid or by incubation with prelabelled cells. A rapid method of extraction was developed using silanised silica cartridges.

Prostaglandin synthesis by glomeruli isolated from rats with glycerol-induced acute renal failure.

In vitro PG synthesis by glomeruli isolated from rats with glycerol-induced acute renal failure (ARF) was measured by radiometric high performance liquid chromatography after incubation with [14C]arachidonic acid and radioimmunoassay (RIA). The four PGs, 6-keto-PGF1 alpha, TXB2, PGF2 alpha, and PGE2 were each synthesized by glomeruli from both control and treated rats but the synthesis rates were greater after glycerol. This increase was not apparent 1 hour after injection but, at 24 hours, all PGs were produced in greater amounts by glomeruli of treated rats. Thus, we studied PGE2, PGE2 alpha, and TXB2 synthesis by glomeruli at various time intervals after induction of ARF using direct RIA, PGF 2 alpha and TXB2 synthesis were greater only at 24 hours and only in the presence of arachidonic acid, whereas PGE2 synthesis was greater at 24 hours, irrespective of arachidonic acid, but at 48 hours only with arachidonic acid. The stimulatory effect of arachidonic acid was always greater in glycerol-treated than in control rats for these three PGs in the later period, whereas a significant decrease for PGE2 was observed at 1 hour. The late increase in PG synthesis may be due to stimulation of the renin-angiotensin system since it was abolished in rats pretreated for 48 hours with captopril. A late increase in PG synthesis by the papilla of the treated rats also was observed. We concluded that any increase in the glomerular production of vasoconstrictor PGs could contribute to the maintenance of acute renal failure, whereas the early fall in the stimulatory effect of arachidonic acid on PGE2 synthesis could play a role in its initiation.