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5 coumarins used in perfumery, cosmetics, therapeutic ointments or occurring naturally were investigated by Freund's complete adjuvant technique (FCAT) in guinea pigs to determine their contact sensitizing potency. 4-Methylesculetin was also studied. Esculetin, dihydrocoumarin and daphnetin were found to be moderate to strong sensitizers, while fraxetin was nearly and 7-methylcoumarin completely inactive. The results corroborate the hypothetical view that only those coumarins having a catecholic disubstitution in the benzene ring, e.g., esculetin, 4-methylesculetin, daphnetin, can become sensitizers on the basis that they are capable of forming ortho-quinones under oxidizing conditions.

From the herbLedum palustre L., collected in July in Kostroma province we have isolated β-sitosterol, the triterpenoids taraxerol, uvaol, and ursolic acid, the coumarin fraxetin, the known flavonoids quercetin, hyperoside, and 3,3′,7-tri-O-methylquercetin, and a new natural flavonoid. The latter was isolated in the form of light yellow crystals with mp 188–190°C (from ethanol), with the composition C19H18O8. On the basis of IR, UV, NMR, and mass spectroscopy the structure of 4′,5-dihydroxy-3,3′,5′,7-tetramethoxyflavone has been suggested for it.

Direct testing of 14 C-labelled aesculetin has yielded results in accord with its previously postulated role as the intermediate between umbelliferone and scopoletin in the biosynthesis of puberulin by A. puberula . Mass spectrometric analysis of an acidic fraction from this species, which should contain all its free phenolic coumarins, indicated the presence of isofraxidin and fraxetin, which was confirmed by GC. [8- O - 14 Me] Isofraxidin was, on average, utilized comparably to [ 14 Me] scopoletin in puberulin formation, and in two experiments the incorporations of > 1% were among the best that have been observed for any puberulin precursor. Participation of isofraxidin strongly implicates fraxetin as an intermediate. The sequence: p -coumaric acid → → umbelliferone → aesculetin → scopoletin → fraxetin → isofraxidin → puberulin is proposed as a feasible biosynthetic route.

Coumaris ofHaplophyllum obtusifolium Ledeb. — obtusinin, capensin, and fraxetin 7-O-β-D-glucopyranoside — have been investigated by13C NMR spectroscopy. Several distinctive features of their NMR spectra have been observed. A complete assignment of the13C NMR spectra of these coumarins has been made and their structures have been confirmed.

A phytochemical investigation of an ethanolic extract of the whole plant of Echites hirsuta (Apocynaceae) resulted in the isolation and identification of the flavonoids naringenin, aromadendrin (dihydrokaempferol), and kaempferol; the coumarin fraxetin; the triterpene ursolic acid; and the sterol glycoside sitosteryl glucoside.

An investigation of the roots of Jatropha glandulifera revealed the presence of a coumarino-lignan, along with jatropholone-A and fraxetin.

The effects of various coumarins (i.e. esculetin, daphnetin and fraxetin) on the formation of the 5-lipoxygenase product, 5-HETE, and the cyclooxygenase product, HHT, were studied. Esculetin (6,7-dihydroxycoumarin) was found to inhibit the formation of 5-HETE more strongly than HHT; its concentrations for 50% inhibition (IC50) were 1.46 +/- 1.02 microM for the formation 5-HETE and 57.3 +/- 17.3 microM for the formation of HHT. Daphnetin (7,8-dihydroxycoumarin) and fraxetin (6-methoxy-7,8-dihydroxycoumarin) also inhibited the formation of the 5-lipoxygenase product, 5-HETE, and the cyclooxygenase product, HHT; their IC50 values were, respectively, 6.90 +/- 2.07 microM and 2.57 +/- 0.088 microM for the formation of 5-HETE and 139.0 +/- 30.0 microM and 532.5 +/- 33.0 microM for the formation of HHT. The monohydroxy coumarin derivatives umbelliferone (7-hydroxycoumarin) and scopoletin (6-methoxy-7-hydroxycoumarin) and the coumarin glucosides fraxin (6-methoxy-7,8-dihydroxycoumarin 8-O-D-glucoside) and esculin (6,7-dihydroxycoumarin 6-O-D-glucoside) also inhibited the formation of 5-HETE, though less strongly. 4-Hydroxycoumarin and coumarin had no effect on either 5-lipoxygenase or cyclooxygenase at concentrations of up to 1 mM. Esculetin inhibited the formation of 5-HETE noncompetitively. In contrast, the dimethoxycoumarin fraxidin (6,8-dimethoxy-7-hydroxycoumarin) inhibited the formation of HHT more strongly than the formation of 5-HETE at a concentration of 1 mM.

It is striking that apart from coumarin, hydrocoumarin, and dicoumarol, all naturally occurring coumarins have an oxygen atom at the 7-position i. e. they can be regarded as derivatives of umbelliferone, which is one of the most widely distributed compounds of this class. The simpler members of the group are hydroxylated or alkoxylated coumarins derived from polyhydric phenols containing a resorcinol system,—a fact which accounts for the prominence of the umbelliferone nucleus; thus aesculetin is derived from hydroxyquinol, daphnetin from pyrogallol, citropten from phloroglucinol, and fraxetin from 1:2:3:4-tetrahydroxybenzene. Hydroxyl groups have not been found in position 3 of the coumarin system, and only two authentic cases (dicoumarol and ammoresinol) are known with hydroxyl in position 4. Frequently the oxygen at site 7 is combined in a third ring which may be fused in either of the two possible positions with respect to the α-pyrone system. Only one case, that of alloxanthoxyletin, is known where any other oxygen atom is concerned in the formation of the third ring (pp. 281).

Fraxetin belongs to an extensive group of natural phenolic antioxidants. We have investigated the modifications in endogenous antioxidant capacity; superoxide dismutase (SOD), catalase (CAT), total and selenium-dependent glutathione peroxidases (GPx) and glutathione reductase (GR) and stress index; glutathione disulphide (GSSG)/reduced glutathione (GSH) ratio and thiobarbituric acid-reactive substances (TBARs) in liver and brain supernatants of C57BL/6J male 12-month-old mice under fraxetin treatment for 30 days. Liver SOD and GPx (total and Se-dependent) activities were not significantly affected by fraxetin, whereas they were increased in the brain compared with control animals. GR activity increased significantly only in the liver of treated mice. Fraxetin treatment-related decreases were shown for GSSG/GSH ratio and rate of accumulation of TBARs (not significant in TBARs) in both tissues. We concluded that the net effect of fraxetin treatment on endogenous antioxidant capacity suggests that this compound might provide an important resistance to, or protection against, free-radical-mediated events which contribute to degenerative diseases of ageing.

A new triterpenoid saponin, 23-acetoxy,16 alpha-hydroxy-13,28-epoxyolean-11-en-3 beta-yl--beta-D-glucopyranosyl- (1-->2)---beta-D-glucopyranosil(1-->3)-beta-D-fucopyranoside, has been isolated from the roots of Bupleurum fruticosum, along with eight coumarins, scopoletin, scoparone, prenyletin, capensin, fraxetin, aesculetin,7-(3-methyl-2-butenyloxy-6-methoxycoumarin, 7-(2-hydroxy-3-methyl-3-butenyloxy)-6-methoxycoumarin and 5,7-dihydroxy-6-methoxy-8-(3-methyl-2-butenyl)coumarin. The last compound has not been previously encountered in nature. The structures were determined by analysis of NMR spectral data including two-dimensional techniques.

We have evaluated the effects of an oral treatment of mice with fraxetin (25 mg/kg for 30 days) on the glutathione system (GSH, GSSG, and GSSG/GSH ratio as stress index), glutathione reductase (GR) and glutathione peroxidase (GPx) in liver supernatants from male C57BL/6J mice (18-month old). A significant antioxidant effect in vivo was found under this treatment by a decrease in the GSSG/GSH ratio and an increased activity of GR compared with the control mice. GSSG rate and GSSG/GSH ratio were correlated with the decline of GPx activity. Our results of increased GR activity could be considered as a supercompensation in glutathione redox status that involves a decrease in the accumulation of GSSG, as well as, in GSSG /GSH ratio. Finally, we suggest that this possible mechanism of supercompensation could lead to an enhancement in the average life span.