Effect of dehydroleucodine on the reproductive tract of male mice.

The effects of a sesquiterpene lactone, dehydroleucodine, on the reproductive tract were investigated using adult male mice. Dehydroleucodine was dissolved in tap water and administered as drinking water for 30 days. All the parameters were compared with a control group that received only vehicle. Animals were killed by decapitation and the trunk blood, the testes and the epididymes were collected. Plasma concentrations of testosterone and oestradiol, and testicular weight and concentration of spermatids did not change by dehydroleucodine. Nevertheless, in epididymal cauda dehydroleucodine treatment caused a diminution in sperm number, a decrease in the amount of tubular fluid and a reduction in the activity of the hydrolytic enzyme N-acetyl-ß-d-glucosaminidase. However, the sperm motility was not altered by dehydroleucodine treatment, although sperm binding to zona-free oocytes increased significantly. These results suggest that dehydroleucodine, which has been implicated in the inhibition of aromatase P450, does not affect the plasma concentration of testosterone and oestradiol or testicular activity, whereas altering several epididymal parameters. The epididymis is thus a more sensitive target for dehydroleucodine action.

Effect of dehydroleucodine on meiosis reinitiation in Bufo arenarum denuded oocytes.

In amphibian oocytes meiosis, the transition from G2 to M phase is regulated by the maturation promoting factor (MPF), a complex of the cyclin-dependent kinase p34/cdc2 and cyclin B. In immature oocytes there is an inactive complex (pre-MPF), in which cdc2 is phosphorylated on both Thr-161 and Thr-14/Tyr-15 residues. The dephosphorylation of Thr-14/Tyr-15 is necessary for the start of MPF activation and it is induced by the activation of cdc25 phosphatase. Late, to complete the activation, a small amount of active MPF induces an auto-amplification loop of MPF stimulation (MPF amplification). Dehydroleucodine (DhL) is a sesquiterpenic lactone that inhibits mammalian cell proliferation in G2. We asked whether DhL interferes with MPF activation. For this question, the effect of DhL (up to 30 microM) on the resumption of meiosis was evaluated, and visualized by germinal vesicle break down (GVBD), of Bufo arenarum oocytes induced in vitro by either: (i) removing follicle cells; (ii) progesterone stimulation; (iii) VG-content injection; or (iv) injection of mature cytoplasm. The results show that DhL induced GVBD inhibition, in a dose-dependent manner, in spontaneous and progesterone-induced oocyte maturation. Nevertheless, DhL at the doses assayed had no effect on GVBD induced by mature cytoplasm injection, but exerted an inhibitory effect on GVBD induced by GV content. On the basis of these results, we interpreted that DhL does not inhibit MPF amplification and that the target of DhL is any event in the early stages of the cdc25 activation cascade.

Effects of Zuccagnia punctata on the gastrointestinal tract in rats and mice.

A pharmacological evaluation of Zuccagnia punctata Cav. (Fabaceae) on the gastrointestinal tract was made in rats and mice. 2',4'-dihydroxychalcone and 2',4'-dihydroxy-3'-methoxychalcone were isolated from Zuccagnia punctata. The data reported in the present work indicate that the acetone extract and infusion of Zuccagnia punctata reduced intestinal transit in rats and mice and offered protection against ethanol-induced ulceration in rats. The Z. punctata effect could be due, in part, to the presence of 2',4'-dihydroxychalcone and 2',4'-dihydroxy-3'-methoxychalcone in this plant.

Sesquiterpene lactone dehydroleucodine selectively induces transient arrest in G2 in Allium cepa root meristematic cells.

Dehydroleucodine is a sesquiterpene lactone recently isolated from aerial parts of a medicinal herb, Artemisia douglasiana Besser. We have previously shown that 25 and 100 microM dehydroleucodine slowed down onion root growth by 30 and 70%, respectively, affecting neither cell viability nor cell elongation. In the present study we analyze the effect of dehydroleucodine on cell cycle phases in onion (Allium cepa L.) root meristematic cells synchronized with caffeine or caffeine and hydroxyurea. Synchronized root cells treated with 100 and 200 microM dehydroleucodine present an interphase lengthening of 5.2 h and 8.2 h, respectively. The S-phase length, estimated by [3H]thymidine incorporation assay, was 6 h for both control roots and roots that had been immersed in dehydroleucodine. The peak of [3H]leucine incorporation was observed 6 h after release from synchronization in controls and in dehydroleucodine-treated roots, indicating that protein synthesis in G2 was not affected. Thus, these results show that dose-dependently dehydroleucodine selectively induces a transient arrest of meristematic cell in G2 and that dehydroleucodine can be used experimentally as a cell cycle suppressor.

The sesquiterpene lactone dehydroleucodine reversibly inhibits Allium cepa L. root growth.

Here, we prove that dehydroleucodine, a sesquiterpene lactone, at low concentrations (25-100 microM) slowed down the Allium cepa L root growth by 22-70% respectively neither affecting cell viability nor cell size. Removal of the drug after 24 h incubation restored the normal growth rate of the roots. Higher concentrations (200 microM) of dehydroleucodine were deleterious for the roots. As cell size did not change, it is most likely that dehydroleucodine affected some event of cell division cycle making it longer. Thus, dehydroleucodine could be a useful tool to slow down cell proliferation.

Two new labdane diterpene glycosides from flowers of Baccharis medullosa DC.

From the flowers of Baccharis medullosa DC (Asteraceae) two new ent-labdane-type diterpene glycosides were isolated. Structures were established by application of various spectroscopic techniques. The sugar residues were identified after acid hydrolysis of the isolated products.

The sesquiterpene lactone dehydroleucodine reversibly inhibits Allium cepa L. root growth.

Here, we prove that dehydroleucodine, a sesquiterpene lactone, at low concentrations (25-100 microM) slowed down the Allium cepa L root growth by 22-70

respectively neither affecting cell viability nor cell size. Removal of the drug after 24 h incubation restored the normal growth rate of the roots. Higher concentrations (200 microM) of dehydroleucodine were deleterious for the roots. As cell size did not change, it is most likely that dehydroleucodine affected some event of cell division cycle making it longer. Thus, dehydroleucodine could be a useful tool to slow down cell proliferation.

Icetexane and abietane diterpenoids from Salvia gilliessi.

One icetexane and two abietane diterpenes were isolated from the aerial parts of Salvia gilliesii, and characterized as 5-epi-icetexone; 12-hydroxy-11,14-diketo-6,8,12-abietatrien-19,20-olide and 6 alpha,12,19-trihydroxy-11,14-diketo-8,12-abietadien-20,7 beta-olide, respectively. The structures were established by analysis of their 1H and 13C NMR spectra with the aid of 2D experiments. The triterpene oleanolic acid was isolated from the same source.

Gastric anti-ulcer activity of several alpha,beta-unsaturated carbonyl compounds in rats.

The gastric cytoprotective activity of several molecules containing an alpha,beta-unsaturated carbonyl system is reported. We attributed this gastroprotective activity to the presence of a non-hindered Michael acceptor in the molecules assayed and suggested that the mechanism of protection would involve, at least in part, a nucleophilic attack of the sulphydryl group of the gastric mucosa to the beta carbon of the Michael acceptors of the compounds assayed.

The sesquiterpene lactone dehydroleucodine (DhL) affects the growth of cultured epimastigotes of Trypanosoma cruzi.

Here, we report an inhibitory effect of a sesquiterpene lactone dehydroleucodine (DhL) on the growth of Trypanosoma cruzi in culture. At concentrations of the drug between 5 and 10 microg/ml in the medium,the parasites remained alive for at least 4 days. Higher concentrations of DhL were lethal for the parasites within a few hours. The effect of DhL is irreversible. Morphological changes induced by DhL were also observed in the parasites. The effect of DhL was blocked by the presence of reducing substrates such as glutathione or dithiothreitol, but these agents were not able to reverse the effect of DhL if added 2 days after the start of drug exposure.

Diterpenes from Laennecia sophiifolia.

From the aerial parts of Laennecia sophiifolia (Kunth) G.L. Nesom, a neo-clerodane and an acyclic furano diterpene were isolated, along with four known compounds, 2beta-hydroxyhardwickiic acid, hawtriwaic acid, apigenin, and beta-sitosterol. Their structures were established as 12-epi-bacchotricuneatin A and (2E,6E)-9-(3-furyl)-6-methyl-2-(4-methylpent-3-enyl)-nona-2,6-dienoic acid, by analysis of spectral evidence. The absolute structure of 12-epi-bacchotricuneatin A was determined by a circular dichroism spectral comparison with that of bacchotricuneatin A.

Effect of dehydroleucodine in experimental colitis in rats and mice.

Dehydroleucodine (DhL), a sesquiterpene lactone (SQL) of the guaianolide type isolated from Artemisia douglasiana Besser, shows a pharmacological cytoprotective effect and significantly prevents the formation of gastric and duodenal lesions induced by various necrotising agents in rodents. The effects of DhL, on two models of experimental colitis were examined. Colitis was produced in male Wistar rats by rectal instillation of 5 and 10% acetic acid, following the methods of Eliakim et al. and Le Duc et al., respectively. In mice colitis was produced by rectal instillation of 0.1 ml of 2,4,6-trinitrobenzene sulphonic acid (5 mg in 50% ethanol) (TNB) as previously described by Chin et al. In this study, the administration of DhL 40 mg kg(-1)(1 h before the induction of colitis) significantly decreased mucosal damage. This effect was consistent in both models. The protection provided by DhL was accompanied by significant decreases in diarrhoea and colon weight; and histologically normal mucosa without ulceration and mucus production were observed. This study shows that both TNB and acetic acid colitis can be pharmacologically controlled by DhL. Our results suggest that the protective activity of DhL in experimental colitis is mediated, at least in part, through the increase of glycoprotein synthesis, anti-inflammatory effect and inhibition of COX-2 induction, and by inhibiting the degranulation of cells containing monoamines.

Growth-inhibitory activities of benzofuran and chromene derivatives toward Tenebrio molitor.

Growth-inhibitory activities of selected natural benzofurans (4-9), trans-cinnamic acid derivatives (10-13), chromene compounds (14 and 16), and some semisynthetic derivatives were determined in last instar larvae of Tenebrio molitor via topical administration in Me2CO. The most inhibitory of the tested compounds were 3-gamma, gamma-dimethylallyl-p-coumaric acid (10) and the benzofuran derivative 12-(p-cumaroyloxy)-tremetone (5), the former compound acting on the pupae and the latter on the last instar larvae. Several developmental deficiencies were observed, and some structure-activity relationships are discussed.

Gastric cytoprotective activity of dehydroleucodine in rats. Role of nitric oxide.

Previously we reported that dehydroleucodine (DhL), a sesquiterpene lactone, shows gastric and duodenal cytoprotective activity. The mechanism is not mediated by antiacid secretory action; DhL stimulated mucus production and indomethacin pretreatment reduced cytoprotective action. In the present study we demonstrated that the gastric cytoprotective effect is antagonized by the nitric oxide (NO) synthase inhibitor, NG-nitro-L-arginine. The inhibitory action of NG-nitro-L-arginine is reversed by L-arginine, but not D-arginine. The findings suggest that NO is involved in the gastroprotection induced by DhL.

Gastric cytoprotective activity of dehydroleucodine in rats. Role of prostaglandins.

Previously, we reported that dehydroleucodine (DhL), a sesquiterpene lactone, protected the gastric mucosa of rats from absolute ethanol-induced lesions in a dose-dependent fashion. The mechanism is not mediated by an antiacid secretory action and DhL stimulated mucous production. In the present study, we report the effect of DhL on the mucosal production of prostaglandin E (PGE) and the mucosal release of PGE2 in rats stomach. DhL in acute treatment does not modify these values decreased by previous treatment with indomethacin or absolute ethanol. However, DhL in subchronic treatment significantly enhanced the mucosal production of PGE and the mucosal release of PGE2. Also, indomethacin pretreatment resulted in a significant reduction of the cytoprotective action of DhL. These results indicate the participation of endogenous prostaglandins in DhL protection against ethanol damage. Moreover, we suggest that the gastric protective activity of DhL against ethanol induced gastric mucosal damage is mediated, at least in part, through PGE and PGE2 in subchronic treatment.

Structure-cytoprotective activity relationship of simple molecules containing an alpha,beta-unsaturated carbonyl system.

In previous reports we attributed the cytoprotective activity of several sesquiterpene lactones to the presence of a nonhindered electrophilic acceptor in their structure. We suggested that the mechanism of protection would be, at least in part, mediated through a reaction between the electrophilic acceptor and the sulfhydryl-containing groups of the mucosa. We report here the gastric cytoprotective effect of simple molecules containing an alpha, beta-unsaturated carbonyl group. In the present paper, we undertake the study of molecular accessibility and molecular shape, in addition to conformational, electronic, and steric factors. Our results helped to establish two important facts connecting chemical structure with cytoprotective effect. Firstly, an adequate molecular accessibility appears to be necessary to produce the biological response, and secondly, the alpha,beta-unsaturated carbonyl system has to be included in a cyclic structure or, at least, in the proximity of a cyclic system.

Gastric cytoprotective activity of 2-cyclopenten-1-one and related compounds.

The cytoprotective activity of the isolated functional groups of several sesquiterpene lactones is reported. Among them the highest activity is shown by alpha-methylen-gamma-butyrolactone and 2-cyclopenten-1-one. The activity shown by those Michael acceptors with a beta carbon hindered by an alkyl substituent was always lower or almost null. A three-way mechanism of action is proposed: a) reduced glutathione synthesis, b) prostaglandin synthesis and c) mucosal glycoprotein synthesis.

Insect antifeedant activity of clerodane diterpenoids.

Fourteen clerodane-type diterpenoids isolated from plants in the genera Baccharis, Teucrium, and Salvia were assayed for antifeedant activity against Tenebrio molitor larvae in order to establish structure-activity relationships. Among the compounds tested, furanoditerpenes with alpha, beta-unsaturated-gamma-lactone moieties, or C-4-epoxy substitution with C-5-methylacetoxy or C-12-acyloxy functionalities, exhibited maximal antifeedant and repellent activities.

Mucus synthesis and sulfhydryl groups in cytoprotection mediated by dehydroleucodine, a sesquiterpene lactone.

The aerial parts of Artemisia douglasiana have been used in folk medicine as a cytoprotective agent against the development of peptic ulcer. Dehydroleucodine [1], its active principle, significantly prevents the formation of gastric lesions induced by the exposure of the rats to absolute ErOH. It was found in this study that (a) pretreatment of rats with 1 (40 mg/kg, po) caused a significant increase in glycoprotein synthetic activity, approximately sevenfold as large as that of the control; and (b) pretreatment of the rats with the thiol reagent N-methylmaleimide (NEM) significantly diminished the cytoprotection provided by 1. However, the protective effect of 1 was not totally abolished by pretreatment with a combination of indomethacin and NEM, indicating additional mechanisms are involved in the cytoprotective action of 1.

Structure-activity relationship in the cytoprotective effect of helenalin and related compounds.

Our previous reports indicate that the cytoprotective effect of helenalin and several sesquiterpene lactones is mediated through a Michael reaction between the sulfhydryl containing peptides of the gastric mucosa and Michael acceptors present in the sesquiterpene lactone molecules. In the present work the different alternative active sites in molecules containing up to three places which could act as viable Michael acceptors are evaluated. To do this, an extensive conformational and electronic study of helenalin and its derivatives was carried out. The experimental and theoretical results obtained show the gamma-lactone and cyclopentenone groups to be the biologically active places of the molecules under study, whereas tiglic and angelic substituents in these compounds show different behavior as Michael acceptors.