Gotthard Wurm

In vitro Inhibition and Stimulation of Purified Prostaglandin Endoperoxide Synthase by Flavonoids: Structure-Activity Relationship

We studied the effects of 37 flavonoids on prostaglandin endoperoxide synthase (EC 1.14.99.1) purified from sheep vesicular glands. Nonplanar flavans were more potent inhibitors than planar flavones and flavonols (IC50 values were, e.g., 40 mumol/l for catechin and epicatechin, 110 mumol/l for galangin, 490 mumol/l for quercetin and 450 mumol/l for kaempherol). Different inhibition mechanisms were observed, i.e. uncompetitive inhibition for nonplanar flavonoids and competitive or noncompetitive inhibition for planar flavonoids. Potent inhibitors in the group of flavones were substances with an o-dihydroxy structure in the B ring and in the group of flavonol substances with two hydroxyl groups in position 5 and 7 of the A ring. None of the flavanones studied caused significant inhibition, except for the flavanone-3-ol, silibinin (silybin), which caused potent inhibition with an IC50 of 120 mumol/l. Several flavonoids, which were able to inhibit the prostaglandin endoperoxide synthase at higher concentrations, were also able to stimulate the enzyme at lower concentrations. These results indicate that the flavonoids should be divided into two groups according to their capacity to inhibit the prostaglandin endoperoxide synthase, represented by planar and nonplanar substances as in each group a close correlation between structure and inhibitory activity was observed.

Interactions of sulfhydryl agents and soybean lipoxygenase inhibitors.

Abstract The enzymic conversion of 5,8,11,14-eicosatetraenoic acid to the corresponding hydroperoxy fatty acids by soybean lipoxygenase (lineolate: oxygen oxidoreductase E.C. 1.13.11.12.) was investigated and a simple selective extraction method was introduced. The known inhibition of the lipoxygenase pathway by phenidone, mercuric chloride, methylmercuric chloride, methylmercuric iodide, 1,5-dihydroxynaphthalene and acetone phenylhydrazone was influenced by thiol compounds in different ways. (1) A total reactivation of lipoxygenase activity was achieved when several thiol compounds, especially gluthatione, were preincubated with the inhibitor mercuric chloride and the enzyme. (2) A remarkable reduction of the inhibitory potency of phenidone against soybean lipoxygenase was seen when thiol compounds were preincubated with the enzyme before the addition of the inhibitor. When phenidone was preincubated with lipoxygenase first, sulfhydryl agents did not restore the enzyme activity. (3) No interaction was seen, when glutathione or other thiol compounds and the lipoxygenase inhibitors 1,5-dihydroxynaphthalene, nordihydroguaiaretic acid and acetone phenylhydrazone were tested against the enzyme. Therefore, we suggest that soybean lipoxygenase inhibitors may act via different modes of action. It is important to study the mechanisms of lipoxygenase inhibitors, since mammalian lipoxygenase and their products are known to be involved in the inflammatory response.

Inhibition of prostaglandin synthetases derived from neuronal and glial cells and rat renal medulla by ortho-, meta- and para-substituted aminophenolic compounds

Acetophenetidines, acetamidophenols, phenetidines and aminophenols substituted in o-, m- or p-position inhibit prostaglandin-synthetases originating from C 1300 mouse neuroblastoma cells (clone N2A), rat astrocytoma cells (clone C 6) and rat renal medulla. Desacetylated compounds were more potent inhibitors than their corresponding acetyl derivatives and many o- and m-analogues were more active than p-substituted structures like paracetamol (p-acetamidophenol) or phenacetin (p-acetophenetidine). When twelve o-, m- or p-aminophenolic test compounds were compared to acetylsalicyclic acid and indomethacin, o-, and p-phenetidine and o-aminophenol were as effective as acetylsalicyclic acid. All aminophenol derivatives which inhibited prostaglandin synthesis suppressed cultured nervous cell and kidney cyclo-oxygenases to similar extents. Our results suggest that aminophenolic drugs are not more effective against prostaglandin-synthetases in the CNS than against those in the periphery.

Substrates for arachidonic acid co-oxidation with peroxidase/hydrogen peroxide: Further evidence for radical intermediates

We tested the ability of a wide variety of organic compounds, including benzene and phenol derivatives, aromatic amines, pyrazoline derivatives and other non-steroidal anti-inflammatory drugs, to act as cosubstrates during the horseradish peroxidase/hydrogen peroxide-mediated oxygenation of arachidonic acid. Structural requirements for drug activation in our system proved to be an aromatic system and ring substitution by an easily oxidizable group. Complementary substituents modified drug activation. Among the phenol derivatives and aromatic amines we found the meta-substituted compounds to be significantly more effective than their ortho- and para-substituted analogues, indicating the involvement of radical intermediates in this type of reaction. The radical from 1-phenyl 3-methyl 2-pyrazolone(5) was detected by electron paramagnetic resonance spectroscopy. Kinetic studies on this radical were in good accordance with time-dependent measurement of arachidonic acid oxygenation.

Decreasing inhibitory potency of prostaglandin synthetase inhibitors during their cooxidative metabolism. Studies on aminophenols, pyrazolon derivatives and 1,3-diphenylisobenzofuran.

A variety of prostaglandin synthetase inhibitors are cooxygenated during arachidonic acid peroxidation catalyzed by rat renal medulla prostaglandin synthetase or soybean lipoxygenase. Phenylbutazone, aminopyrine, 1,3-diphenylisobenzofuran, paracetamol, p-aminophenol, p-phenetidine and other o- and m-substituted aminophenol derivatives were cooxygenated, whereby prostaglandin synthetase inhibition was significantly weakened due to the formation of less inhibitory metabolites. In contrast, the inhibitory potency of diclofenac, indomethacin and phenacetin and its analogues remained unchanged during prostaglandin synthesis inhibition, because these compounds were no suitable cooxygenation substrates. Evidence is given that quinone imines may not be involved in the cooxidative metabolism of paracetamol and other aminophenols. As to the mechanisms of cooxygenation of suitable substrates dependent on their chemical structures either the arachidonic acid oxygenase or the subsequent hydroperoxidase reaction may trigger the oxygenation. 1,3-Diphenylisobenzofuran is metabolized during the formation of arachidonic acid hydroperoxides in contrast to paracetamol, which requires an additional peroxidase reaction to yield reactive metabolites.