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Kaurane diterpenes are considered important compounds in the development of new highly effective anticancer chemotherapeutic agents. Genotoxic effects of anticancer drugs in non-tumour cells are of special significance due to the possibility that they induce secondary tumours in cancer patients. In this context, we evaluated the genotoxic and mutagenic potential of the natural diterpenoid kaurenoic acid (KA), i.e. (-)-kaur-16-en-19-oic acid, isolated from Xylopia sericeae St. Hill, using several standard in vitro and in vivo protocols (comet, chromosomal aberration, micronucleus and Saccharomyces cerevisiae assays). Also, an analysis of structure-activity relationships was performed with two natural diterpenoid compounds, 14-hydroxy-kaurane (1) and xylopic acid (2), isolated from X. sericeae, and three semi-synthetic derivatives of KA (3-5). In addition, considering the importance of the exocyclic double bond (C16) moiety as an active pharmacophore of KA cytotoxicity, we also evaluated the hydrogenated derivative of KA, (-)-kauran-19-oic acid (KAH), to determine the role of the exocyclic bond (C16) in the genotoxic activity of KA. In summary, the present study shows that KA is genotoxic and mutagenic in human peripheral blood leukocytes (PBLs), yeast (S. cerevisiae) and mice (bone marrow, liver and kidney) probably due to the generation of DNA double-strand breaks (DSB) and/or inhibition of topoisomerase I. Unlike KA, compounds 1-5 and KAH are completely devoid of genotoxic and mutagenic effects under the experimental conditions used in this study, suggesting that the exocyclic double bond (C16) moiety may be the active pharmacophore of the genetic toxicity of KA., (2010 Elsevier B.V. All rights reserved.)

Copaiba oil extracted from the Amazon traditional medicinal plant Copaifera langsdorffii is rich in kaurenoic acid (ent-kaur-16-en-19-oic acid), a diterpene that has been shown to exert anti-inflammatory, hypotensive, and diuretic effects in vivo and antimicrobial, smooth muscle relaxant and cytotoxic actions in vitro. This study evaluated its potential genotoxicity against Chinese hamster lung fibroblast (V79) cells in vitro, using the Comet and the micronucleus assays. Kaurenoic acid was tested at concentrations of 2.5, 5,10, 30 and 60 microg/mL. The positive control was the methylmethanesulfonate (MMS). The duration of the treatment of V79 cells with these agents was 3h. The results showed that unlike MMS, kaurenoic acid (2.5, 5, and 10 microg/mL) failed to induce significantly elevated cell DNA damage or the micronucleus frequencies in the studied tests. However, exposure of V79 cells to higher concentrations of kaurenoic acid (30 and 60 microg/mL) caused significant increases in cell damage index and frequency. The data obtained provide support to the view that the diterpene kaurenoic acid induces genotoxicity.

The effect of kaurenoic acid, a diterpene isolated from the oleo-resin of the popular medicinal plant Copaifera langsdorffii (Leguminaceae), was analysed on rat uterine muscle responsiveness to various drugs in vitro. Cumulative concentration-response curves to acetylcholine and oxytocin were obtained before and after incubation of uterine segments with up to 160 microm of kaurenoic acid. The maximal contractile response (E(max)) evoked by these agonists was inhibited by kaurenoic acid in a concentration-related manner; at 160 microm, kaurenoic acid depressed the E(max) of oxytocin and acetylcholine by 83% and 91%, respectively. The relaxation caused by kaurenoic acid on oxytocin-induced contraction was unaffected in the presence of tetraethyl ammonium, a compound that blocks the calcium activated potassium channels. It was partially reversed by glibenclamide (10(-5) m), an ATP-sensitive potassium channel blocker. Also, kaurenoic acid at 160 microm concentration was found to inhibit significantly the CaCl(2)-evoked contractile responses in a medium of high potassium and zero calcium. Furthermore, kaurenoic acid was found to relax the sustained tonic contraction induced by acetylcholine, oxytocin, BaCl(2) and KCl in a concentration-dependent way. However, KCl-induced tonic contraction was only weakly inhibited by kaurenoic acid. These data indicate that the diterpene, kaurenoic acid, exerts a uterine relaxant effect acting principally through calcium blockade and in part, by the opening of ATP-sensitive potassium channels., (Copyright 2003 John Wiley & Sons, Ltd.)

Kaurenoic acid, a diterpene from Copaifera langsdorffii (Leguminaceae), was evaluated on rat colitis induced by acetic acid. Rats were pretreated orally (15 and 2 h before) or rectally 2 h before induction of colitis with kaurenoic acid (50 and 100 mg/kg) or vehicle (1 ml, 3% DMSO). Colitis was induced by intracolonic instillation of a 2 ml of 4% (v/v) acetic acid solution and, 24 h later, the colonic mucosal damage was analysed macroscopically for the severity of mucosal damage, the myeloperoxidase (MPO) activity and the malondialdehyde (MDA) levels in the colon segments. A marked reduction in gross damage score (52% and 42%) and wet weight of damaged colon tissue (39% and 32%) were observed in rats that received 100 mg/kg kaurenoic acid, respectively, by rectal and oral routes. This effect was confirmed biochemically by a two- to three-fold reduction of colitis associated increase in MPO activity, the marker of neutrophilic infiltration and by a marked decrease in MDA level, an indicator of lipoperoxidation in colon tissue. Furthermore, light microscopy revealed the marked diminution of inflammatory cell infiltration and submucosal edema formation in the colon segments of rats treated with the test compound. These findings indicate the anti-inflammatory potential of kaurenoic acid in acetic acid-induced colitis.

In this work, we studied the effects of kaurenoic acid, a diterpene isolated from the oleo-resin of Copaifera langsdorffii in developing sea urchin (Lytechinus variegatus) embryos, on tumor cell growth in microculture tetrazolium (MTT) test and on mouse and human erythrocytes in hemolysis assay. Continuous exposure of embryos to kaurenoic acid starting immediately after fertilization inhibited the first cleavage (IC(50): 84.2 microM) and progressively induced embryo destruction (IC(50): 44.7 microM and < 10 microM for blastulae and larvae stages, respectively). In MTT assay, kaurenoic acid at a concentration of 78 microM produced growth inhibition of CEM leukemic cells by 95%, MCF-7 breast and HCT-8 colon cancer cells by 45% each. Further, kaurenoic acid induced a dose-dependent hemolysis of mouse and human erythrocytes with an EC(50) of 74.0 and 56.4 microM, respectively. The destruction of sea urchin embryos, the inhibition of tumor cell growth and the hemolysis of mouse and human erythrocytes indicate the potential cytotoxicity of kaurenoic acid.

Aim: The objective of this study was to assess in rats the antidiabetic effects (i.e. reduction of hyperglycaemia and hypertriglyceridaemia) of trans-dehydrocrotonin (t-DCTN), a bioactive diterpene isolated from the popular medicinal plant Croton cajucara., Methods: Hyperglycaemia was induced by intraperitoneal injection of streptozotocin (STZ) and hypertriglyceridaemia by oral administration of ethanol in rats. The blood glucose levels were measured by the glucose oxidase method using commercially available enzyme kits., Results: Treating rats with t-DCTN (50 mg/kg) significantly reduced STZ-induced increases in blood glucose levels as well as ethanol-induced increases in blood triglycerides., Conclusion: The results suggest that t-DCTN has an antidiabetic potential that warrants further research on its mechanism and clinical significance.

The effect of trans-dehydrocrotonin (t-DCTN) from Croton cajucara Benth. was investigated in mice on Triton WR 1339 (tyloxapol)-induced hypercholesterolaemia and hypertriglyceridaemia. Mice treated with single application of tyloxapol (400 mg/kg, i.p.) demonstrated significantly increased blood levels of total cholesterol and triglycerides at 24 h and 48 h after its injection, compared to normal controls. These increases were found to be markedly suppressed in animals treated orally with 25 and 50 mg/kg t-DCTN or 100 mg/kg gemfibrozil, an established antihypercholesterolaemic drug. These results suggest that t-DCTN may be a suitable candidate for combating pathologies associated with hyperlipaemia.

The clerodane diterpene trans-dehydrocrotonin extracted and isolated from the stem bark of Croton cajucara Benth. was investigated for its lipid-lowering effect in mice fed on a high-fat diet. Mice fed on a high-fat diet for a two-week period demonstrated significantly increased blood levels of total cholesterol and triglycerides, compared with normal controls. Oral treatment with trans-dehydrocrotonin at a dose of 25 or 50 mg kg(-1) daily markedly suppressed the high-fat-diet associated rise in total cholesterol and triglyceride levels. The hypocholesterolaemic effect of trans-dehydrocrotonin was more prominent at the dose of 50 mg kg(-1) with significant decreases in high-density lipoprotein, very-low-density lipoprotein and low-density lipoprotein cholesterol levels. The lower atherogenic index of the trans-dehydrocrotonin-treated groups suggests the hypolipidaemic potential of this plant-based drug. These results indicate that orally administered trans-dehydrocrotonin is effective in suppressing high-fat-diet-induced hyperlipidaemia in mice and suggest its likely beneficial use as anti-atherogenic agent.

This study examined trans-dehydrocrotonin (t-DCTN), a nor-clerodane diterpene isolated from the Brazilian medicinal plant Croton cajucara Benth., for a possible antioestrogenic activity using immature rats as a model system for bioassay of oestrogen, and for an antiimplantation effect in regularly cycling rats of proven fertility. In the antioestrogen test, t-DCTN (25 and 50 mg/kg) effectively prevented oestrogen-induced increases of uterine wet weights. In addition, the vaginal openings provoked by oestrogen were completely prevented by t-DCTN. However, blastocyst-implantation was only insignificantly affected in t-DCTN pretreated animals. These results suggest that t-DCTN may be an antioestrogen and warrants further studies with regard to its mechanism of action., (Copyright 1999 John Wiley & Sons, Ltd.)

trans-Dehydrocrotonin (t-DCTN), a 19-nor-clerodane diterpene isolated from the bark of Croton cajucara Benth. (Euphorbiaceae) demonstrated a significant hypoglycemic activity in alloxan-induced diabetic rats but not in normal rats, at oral doses of 25 and 50 mg/kg body weight. The drug also effectively lowered the blood sugar levels in glucose fed normal rats. The hypoglycemic effect of t-DCTN was almost comparable to that produced by glibenclamide (2 mg/kg), a clinically useful drug. The results indicate the antihyperglycemic potential of t-DCTN.