Ent-kauren-19-oic acid derivatives from the stem bark of Croton pseudopulchellus Pax

Two new ent-kauren-19-oic acid derivatives, ent-14S*-hydroxykaur-16-en-19-oic acid and ent-14S*,17-dihydroxykaur-15-en-19-oic acid together with eleven known compounds ent-kaur-16-en-19-oic acid, ent-kaur-16-en-19-al, ent-12β-hydroxykaur-16-en-19-oic acid, ent-12β-acetoxykaur-16-en-19-oic acid, 8R,13R-epoxylabd-14-ene, eudesm-4(15)-ene-1β,6α-diol, (?)-7-epivaleran-4-one, germacra-4(15), 5E,10(14)-trien-9β-ol, acetyl aleuritolic acid, β-amyrin, and stigmasterol were isolated from the stem bark of Croton pseudopulchellus (Euphorbiaceae). Structures were determined using spectroscopic techniques. Ent-14S*-hydroxykaur-16-en-19-oic acid, ent-kaur-16-en-19-oic acid, ent-12β-hydroxykaur-16-en-19-oic acid, ent-12β-acetoxykaur-16-en-19-oic acid and 8R,13R-epoxylabd-14-ene were tested for their effects on Semliki Forest virus replication and for cytotoxicity against human liver tumour cells (Huh-7 strain) but were found to be inactive. Ent-kaur-16-en-19-oic acid, the major constituent, showed weak activity against the Plasmodium falciparum (CQS) D10 strain.     

Kolavane and kaurane diterpenes from the stem bark of Xylopia aethiopica

The stem bark of Xylopia aethiopica has yielded four diterpenes, two of them novel. Three of the diterpenes were identified as (?)-kaur-16-en-19-oic acid and its 7-oxo and 7β-hydroxy derivatives. The fourth was the novel kolavane derivative 2-oxo-kolav-3,13-dien-15-oic acid, a type of compound not previously recorded in the Annonaceae.     

Terpenoids and a flavan-3-ol from viguiera quinqueradiata

The isolation is reported of the new natural products from Viguiera quinqueradiata, acetylleptocarpin and (2R,3S-4′-hydroxy-3′,5,7-tri-O-methyl-flavan-3-ol. The diterpenes 15α-angeloyloxy-ent-kaur-16-en-19-oic acid, 15α-tigloyloxy-ent-kaur-16-en-19-oic acid and the sesquiterpene lactones leptocarpin and budlein A were also found.     

Diterpenoids and a sesquiterpene lactone from viguiera ladibractate

Eight known diterpene acids, ent-12-oxokaur-9(11),16-dien-19-oic acid, ent-12β-hydroxykaur-9(11),16-dien-19-oic acid, ent-isokaur-15(16)-en-17,19-dioic acid, ent-15α,16-epoxy-17-hydroxykaura-19-oic acid, ent-kaura-17,19-dioic acid, ent-kaur-16-en-19-oic acid, grandifloric acid, angeloyloxygrandifloric acid, as well as a new sesquiterpene lactone, ladibranolide, were isolated from Viguiera ladibractate. The stereochemistry of the sesquiterpene lactone was established by NOE experiments.     

Sesquiterpene lactones and diterpenoids from Helianthus argophyllus

Three germacranolide sesquitepene lactones (argophyllin-A and -B and eupatolide), three diterpenoids (ciliaric acid, (?)-16-α-hydroxy-kaur-11-en-19-oic acid and (?)-16-α-hydroxykaurane) and one flavonoid (nevadensin) were isolated and characterized from a chloroform extract of Helianthus argophyllus. Argophyllin-A and -B are both described here for the first time. Their structures were deduced by ¹H NMR and ¹³C NMR. Argophyllin-A and -B were found to show anti-auxin effects while eupatolide exhibited weak insecticidal activity.     

The biosynthesis of kaurenolide diterpenoids by gibberella fujikuroi

The biosynthesis of 7β-hydroxy- and 7β,18-dihydroxy-kaurenolides from ent-kaur-16-en-19-oic acid has been investigated by incubating unlabelled     

Diterpenoid Total Synthesis

Total syntheses of (±)-steviol (IIa) and (±)-kaur-16-en-19-oic acid (III) were accomplished. A synthesis of methyl (±)-8α-carboxymethylpodocarpan-13-on-19-oate (IVa), a degradation product of steviol, was also described.     

Ent-kaurane derivatives from the root cortex of yacon and other three Smallanthus species (Heliantheae,Asteraceae)

The metabolites produced by the secretory canals of the root cortex from four Smallanthus species belonging to the yacon group were identified as ent-kaurane-type diterpenes. The dichloromethane root cortex extracts of the four species were treated with diazomethane and analyzed comparatively by GC–MS using a simple and rapid procedure which is very sensitive and reproducible permitting detection of minor components. In all cases, ent-16-kauren-19-oic acid (kaurenoic acid) methyl ester was the main component, differences being observed only in the minor components. The minor components identified were grandiflorenic acid methyl ester, ent-16-kauren-19-al, 16α,17-epoxy-15α-angeloyloxy-kauran-19-oic acid methyl ester and several O-acyl derivatives at C-15 or C-18 of kaurenoic acid. One of the minor components, 18-isobutyroyloxy-ent-kaur-16-en-19-oic acid is a new kaurenoic acid derivative. Grandiflorenic acid and 15-α-angeloyloxy-16,17-α-epoxy-ent-16-kauren-19-oic acid were present only in Smallanthus sonchifolius and Smallanthus siegesbeckius which showed very similar GC traces. The different GC profile of RC diterpenes from Smallanthus connatus and Smallanthus macroscyphus supports the view that they are different taxa. Some chemotaxonomic aspects of the genus Smallanthus and the subtribe Milleriinae are briefly discussed.     

Gummiferolic acid,a new ent-atis-16-ene diterpenoid from Margotia gummifera

From the roots of Margotia gummifera a new diterpenoid with the ent-atis-16-ene skeleton, gummiferolic acid, has been obtained in very high yield (2% of the dry plant), together with the known ent-kaur-16-en-19-oic acid.     

Structures of the novel α-glucosyl linked diterpene glycosides from Stevia rebaudiana

From the commercial extract of the leaves of Stevia rebaudiana, two new minor diterpene glycosides having α-glucosyl linkage were isolated besides the known steviol glycosides including stevioside, steviolbioside, rebaudiosides A–F, rubusoside and dulcoside A. The structures of the two compounds were identified as 13-[(2-O-(3-α-O-d-glucopyranosyl)-β-d-glucopyranosyl-3-O-β-d-glucopyranosyl-β-d-glucopyranosyl)oxy] ent-kaur-16-en-19-oic acid β-d-glucopyranosyl ester (1), and 13-[(2-O-β-d-glucopyranosyl-3-O-(4-O-α-d-glucopyranosyl)-β-d-glucopyranosyl-β-d-glucopyranosyl)oxy] ent-kaur-16-en-19-oic acid β-d-glucopyranosyl ester (2), on the basis of extensive NMR and MS spectral data as well as chemical studies.     

Schistosomicidal activity of kaurane,labdane and clerodane-type diterpenes obtained by fungal transformation

Schistosomiasis continues to be a huge challenge for researchers, pharmaceutical companies, and governments in developing countries. Diterpene compounds are good source for the development of novel potential leading compounds to treat schistosomiasis. We are reporting herein the schistosomicidal activity of ent-kaurenoic acid, ent-copalic acid, ent-hardwickiic acid, isolated from oleoresins of Copaifera spp, and of their derivatives obtained by fungal transformation with strains of Aspergillus fumigatus, A. terreus, A. phoenicis and A. ochraceus, and of Cunninghamella echinulata e C. elegans. The in vitro antiparasitical assays were performed using adult worm pairs of Schistosoma mansoni for the evaluation of the worm pairing, egg production, and eggs development. Ten kaurane, labdane and clerodane-type diterpenes were obtained by fungal transformation and 7α-acetoxy-ent-kaur-16-en-19-oic acid and ent-hardwickiic acid were the most active ones by causing mortality of 100 % of the parasites within 24 h (concentrations of 100.0 and 200.0 μM) and displaying respective IC₅₀ values for 24, 48 and 72 h of 56.7, 37.6 and 29.2 μM and 29.6, 30.8 and 25.7 μM. Additionally, 7α-acetoxy-ent-kaur-16-en-19-oic acid and ent-hardwickiic acid highly reduced the number of laid eggs at 6.25 and 12.5 μM, respectively. These diterpenes should be further investigated as potential candidates for antiparasitic drug discovery.     

Ent-kaurenic Acid and Its Related Compounds from Glandular Trichome Exudate and Leaf Extracts of Polymnia sonchifolia

Yacon (Polymnia sonchifolia) leaves possess glandular trichomes on the surface. The exudate from the glandular trichome and the leaf are itself rich in ent-kaurenic acid (ent-kaur-16-en-19-oic acid). A kaurene derivative, 15-α-angeloyloxy-ent-kauren-19-oic acid 16-epoxide, was isolated from the leaves, together with two known angeloyloxykaurenic acids. The high content of ent-kaurenic acid in the leaf suggests that these diterpenes play a certain physiological role, since the glandular trichome exudates of other species function in their defensive mechanism.     

Biotransformation of ent-kaur-16-ene and ent-trachylobane 7β-acetoxy derivatives by the fungus Gibberella fujikuroi (Fusarium fujikuroi)

Candol A (7β-hydroxy-ent-kaur-16-ene) (6) is efficiently transformed by Gibberella fujikuroi into the gibberellin plant hormones. In this work, the biotransformation of its acetate by this fungus has led to the formation of 7β-acetoxy-ent-kaur-16-en-19-oic acid (3), whose corresponding alcohol is a short-lived intermediate in the biosynthesis of gibberellins and seco-ring ent-kaurenoids in this fungus. Further biotransformation of this compound led to the hydroxylation of the 3β-positions to give 7β-acetoxy-3β-hydroxy-ent-kaur-16-en-19-oic acid (14), followed by a 2β- or 18-hydroxylation of this metabolite. The incubation of epicandicandiol 7β-monoacetate (7β-acetoxy-18-hydroxy-ent-kaur-16-ene) (10) produces also the 19-hydroxylation to form the 18,19 diol (20), which is oxidized to give the corresponding C-18 or C-19 acids. These results indicated that the presence of a 7β-acetoxy group does not inhibit the fungal oxidation of C-19 in 7β-acetoxy-ent-kaur-16-ene, but avoids the ring B contraction that leads to the gibberellins and the 6β-hydroxylation necessary for the formation of seco-ring B ent-kaurenoids. The biotransformation of 7β-acetoxy-ent-trachylobane (trachinol acetate) (27) only led to the formation of 7β-acetoxy-18-hydroxy-ent-trachylobane (33).     

Biotransformation of 7α-hydroxy- and 7-oxo-ent-atis-16-ene derivatives by the fungus Gibberella fujikuroi

The microbiological transformation of 7α,19-dihydroxy-ent-atis-16-ene by the fungus Gibberella fujikuroi gave 19-hydroxy-7-oxo-ent-atis-16-ene, 13(R),19-dihydroxy-7-oxo-ent-atis-16-ene, 7α,11β,19-trihydroxy-ent-atis-16-ene and 7α,16β,19-trihydroxy-ent-atis-16-ene, while the incubation of 19-hydroxy-7-oxo-ent-atis-16-ene afforded 13(R),19-dihydroxy-7-oxo-ent-atis-16-ene and 16β,17-dihydroxy-7-oxo-ent-atisan-19-al. The biotransformation of 7-oxo-ent-atis-16-en-19-oic acid gave 6β-hydroxy-7-oxo-ent-atis-16-en-19-oic acid, 6β,16β,17-trihydroxy-7-oxo-19-nor-ent-atis-4(18)-ene and 3β,7α-dihydroxy-6-oxo-ent-atis-16-en-19-oic acid.     

Metabolic transformations of some ent-kaurenes in Gibberella fujikuroi

The conversion of ent-kaur-16-enes to gibberellic acid in Gibberella fujikuroi is blocked by A-ring modifications. Thus ent-3β-hydroxykaur-16-en-19-yl succinate gives good conversion (46%) to the 7β-hydroxy derivative.* Under the same conditions the 3β-epimer gives 7β- or 6α-hydroxylation and the former occurs for the 3-oxo analogue. The succinoyloxy function acts as a less efficient block and ent-kaur-16-en-19-yl succinate is converted to 7β-hydroxy and 6β,7β-dihydroxy derivatives along with gibberellic acid. Hydrolysis of the succinate block of the metabolities provides the 7β, 19-diol and 6β,7β, 19-triol. Of this pair only the former was effectively metabolized to gibberellic acid in G. fujikuroi.     

Structures of the novel diterpene glycosides from Stevia rebaudiana

From the commercial extract of the leaves of Stevia rebaudiana, two new diterpenoid glycosides were isolated besides the known steviol glycosides including stevioside, rebaudiosides A–F, rubusoside, and dulcoside A. The structures of the two new compounds were identified as 13-[(2-O-6-deoxy-β-d-glucopyranosyl-β-d-glucopyranosyl)oxy] ent-kaur-16-en-19-oic acid β-d-glucopyranosyl ester (1), and 13-[(2-O-6-deoxy-β-d-glucopyranosyl-3-O-β-d-glucopyranosyl-β-d-glucopyranosyl)oxy] ent-kaur-16-en-19-oic acid β-d-glucopyranosyl ester (2), on the basis of extensive NMR and MS spectral data as well as chemical studies.     

Tetrachyrin,a new rearranged kaurenoid lactone,and diterpene acids from Tetrachyron orizabaensis and Helianthus debilis

(?)-Kaur-16-en-19-oic acid and tetrachyrin, a new rearranged kaurenoid lactone, were isolated from Tetrachyron orizabaensis var. websteri and Helianthus debilis ssp. debilis. The latter species also afforded angeloylgrandifloric acid.     

Studies on the Specificity and Site of Action of alpha-Cyclopropyl-alpha-[p-methoxyphenyl]-5-pyrimidine Methyl Alcohol (Ancymidol), a Plant Growth Regulator

α-Cyclopropyl-α-[p-methoxyphenyl]-5-pyrimidine methyl alcohol (ancymidol) is an inhibitor of ent-kaur-16-ene oxidation in microsomal preparations from the liquid endosperm of immature Marah macrocarpus seeds. The K_i for this inhibitor is about 2 × 10⁻⁹ m. Ancymidol also blocks ent-kaur-16-en-19-ol and ent-kaur-16-en-19-al oxidation by the same preparations with a similar efficiency, but does not significantly inhibit ent-kaur-16-en-19-oic acid oxidation. Ancymidol appears to be specific for this series of oxidations in higher plant tissues. It does not inhibit the oxidation of kaurene nor kaurenoic acid in rat liver microsomes and has no significant effect on the oxidation of cinnamic acid in microsomal preparations from Sorghum bicolor seedlings. Ancymidol also does not inhibit kaurene oxidation in vitro nor in vivo in cultures of the fungus Fusarium moniliforme. The presence of ancymidol did not significantly alter the activities of NADPH-cytochrome c reductase, NADH-cytochrome c reductase, or NADH-cytochrome b₅ reductase. The addition of ancymidol to suspensions of oxidized M. macrocarpus endosperm led to a difference spectrum with an absorption maximum at 427 nm and a minimum at 410 nm.     

Isolation of gibberellin precursors from heavily pigmented tissues

The kauranoid precursors of gibberellins are difficult to isolate from heavily pigmented plant tissues. In this paper, we describe relatively simple and efficient procedures for the purification of these compounds from tissues containing chlorophyll and other high molecular weight pigments. Extracts of shoots from Thlaspi arvense L. were subjected first to size exclusion chromatography using ethyl acetate as the eluting solvent. This procedure resulted in the separation of kauranoids as a class of compounds from chlorophyll. Typically, a 90% reduction in mass of the kauranoid enriched-fraction was observed. This fraction was subjected to reverse phase high performance liquid chromatography and individual fractions analyzed by combined gas chromatography-mass spectrometry. Five kauranoids were identified in shoot extracts of T. arvense: ent-kaur-16-ene, ent-kaur-16-en-19-ol, ent-kaur-16-en-19-oic acid, trachylobanoic acid, and 7β, 13-dihydroxykaurenolide. The metabolic relationships of these compounds to the gibberellins previously identified in this species (JD Metzger, MC Mardaus [1986] Plant Physiol 80: 396-402) are discussed. In addition, the utility of size exclusion chromatography in preparative situations is demonstrated by the purification of ent-kaurenoic acid in milligram quantities from the florets of Helianthus annuus L.     

Genetics and gibberellin production inGibberella fujikuroi

Gibberella fujikuroi (Fusarium moniliforme) is a complex group of plant pathogens. Some strains produce gibberellic acid and other gibberellins that promote growth and regulate various stages in plant development.The paper describes the research effort directed to development of genetic tools for this species. Furthermore the main features of the gibberellin biosynthetic pathway as established in Gibberella are described.Abbreviations AMO 1618 2-isopropyl-4-(trimethylammonium chloride)-5-methylphenylpiperidine-1-carboxylate - hydroxykaurenoic acid ent-kaur-16-en-7-ol-19-oic acid - kaurenal ent-kaur-16-en-19-al - kaurene ent-kaur-16-ene - kaurenoic acid ent-kaur-16-en-19-oic acid - kaurenol ent-kaur-16-en-19-ol - paclobutrazol 1-(4-chlorophenyl)-4,4-dimethyl-2-(1,2,4-triazol-lyl)pentan-3-ol - pefurazoate pent-4-enyl-N-furfuryl-N-imidazol-1-ylcarbonyl-DL-homoa laninate - tetcyclacis 5-(4-chlorophenyl)-3,4,5,9,10-pentaazatetracyclo-5,4,10^2.6,O^8.11-dodeca-3,9-diene - triarimol -(2,4-dichlorophenyl)--phenyl-5-pyrimidine methyl alcohol