Microbial transformation of cadina-4,10(15)-dien-3-one, aromadendr-1(10)-en-9-one and methyl ursolate by Mucor plumbeus ATCC 4740

The sesquiterpenes cadina-4,10(15)-dien-3-one (1) and aromadendr-1(10)-en-9-one (squamulosone) (14) along with the triterpenoid methyl ursolate (21) were incubated with the fungus Mucor plumbeus ATCC 4740. Substrates 1, 14 and ursolic acid (20) were isolated from the plant Hyptis verticillata in large quantities. M. plumbeus hydroxylated 1 at C-12 and C-14. When the iron content of the medium was reduced, however, hydroxylation at these positions was also accompanied by epoxidation of the exocyclic double bond. In total nine new oxygenated cadinanes have been obtained. Sesquiterpene 14 was converted to the novel 2alpha,13-dihydroxy derivative along with four other metabolites. Methyl ursolate (21) was transformed to a new compound, methyl 3beta,7beta,21beta-trihydroxyursa-9(11),12-dien-28-oate (22). Two other triterpenoids, 3beta,28-dihydroxyurs-12-ene (uvaol) (23) and 3beta,28-bis(dimethylcarbamoxy)urs-12-ene (24) were not transformed by the micro-organism, however.     

Biotransformation of cadinane sesquiterpenes by Beauveria bassiana ATCC 7159

Incubation of cadina-4,10(15)-dien-3-one with Beauveria bassiana ATCC 7159 has resulted in the production of nine novel sesquiterpenes. These metabolites were identified as (4S)-cadin-10(15)-en-3-one, (4S)-3 alpha-hydroxycadin-10(15)-ene, (4R)-3 alpha-hydroxycadin-10(15)-ene, (4S)-3 beta-hydroxycadin-10(15)-ene, (4S)-3 beta-hydroxycadina-10(15),12(14)-diene, (4S)-13-hydroxycadin-10(15)-en-3-one, (4S)-12-hydroxycadin-10(15)-en-3-one, (4R)-3 beta, 14-dihydroxycadin-10(15)-ene and 3 alpha-hydroxycadina-4,10(15)-diene. The allylic alcohol 3 alpha-hydroxycadina-4,10(15)-diene was also biotransformed to afford cadina-4,10(15)-dien-3-one, (4S)-cadin-10(15)-en-3-one and (4S)-12-hydroxycadin-10(15)-en-3-one. The insecticidal potential and phytotoxicity of the isolated metabolites have been evaluated.     

Practical one-pot conversion of 17beta-estradiol to 10beta-hydroxy- (p-quinol) and 10beta-chloro-17beta-hydroxyestra-1,4-dien-3-one

An efficient one-pot procedure for the preparation of 10beta,17beta-dihydroxyestra-1,4-dien-3-one (p-quinol, 1, 75%) is reported, involving oxidation of 17beta-estradiol with potassium permanganate. Similar treatment of 17beta-estradiol with sodium chlorite led to 10beta-chloro-17beta-hydroxyestra-1,4-dien-3-one (2) in 44% yield along with smaller amounts 4-chloro-10beta,17beta-dihydroxyestra-1,4-dien-3-one (3), 2,10beta-dichloro-17beta-hydroxyestra-1,4-dien-3-one (4), and 4,10beta-dichloro-17beta-hydroxyestra-1,4-dien-3-one (5).     

ent-Clerodane diterpenes and other constituents from the liverwort Adelanthus lindenbergianus (Lehm.) Mitt

Eleven ent-clerodanes, 13-hydroxy-cis-ent-cleroda-3,14-diene, 15-hydroxy-cis-ent-cleroda-3,13(E)-diene, 1beta,12:15,16-diepoxy-cis-ent-cleroda-13(16),14-dien-18alpha,6alpha-olide, 8beta,12:15, 16-diepoxy-cis-ent-cleroda-13(16),14-dien-18alpha,6alpha-olide, 1beta,16:15,16-diepoxy-cis-ent-cleroda-12,14-dien-18alpha,6alpha-olide, 7beta,12:8beta,12-diepoxy-15-hydroxy-cis-ent-cleroda-13-en-16,15:18alpha,6alpha-diolide, 7beta,12:8beta,12-diepoxy-16-hydroxy-cis-ent-cleroda-13-en-15,16:18alpha,6alpha-diolide, 1alpha-acetoxy-8beta,12-epoxy-15-hydroxy-cis-ent-cleroda-13-en-16,15:18alpha,6alpha-diolide, 1beta,12-epoxy-16-hydroxy-cis-ent-cleroda-13-en-15,16:18alpha,6alpha-diolide, 8beta,12-epoxy-15-hydroxy-trans-cleroda-13-en-16,15:18alpha,6alpha-diolide, 8beta,12-epoxy-16-hydroxy-trans-cleroda-13-en-15,16:18alpha,6alpha-diolide along with the known clerodane diterpenes anastreptin and orcadensin have been isolated from the liverwort Adelanthus lindenbergianus (Lehm.) Mitt. Furthermore, three eudesmane sesquiterpenes together with the known (-)-1beta,10-epoxyaristolan, 3,4-seco-4(23),20(29)-lupadien-3,28-dicarboxylic acid dimethyl ester and two acetophenone derivatives were identified by spectroscopic methods, essentially MS and NMR experiments.     

New 2(3-->20)abeotaxane and 3,11-cyclotaxane from needles of Taxus cuspidata

Two new taxoid metabolites, 2alpha,7beta,10beta-triacetoxy-5alpha,13alpha-dihydroxy-2(3-->20)abeotaxa-4(20),11-dien-9-one (1) and 2alpha-acetoxy-5alpha-cinnamoyloxy-9alpha,10beta-dihydroxy-3,11-cyclotax-4(20)-en-13-one (2), were isolated from the methanol extract of needles of the Japanese yew, Taxus cuspidata.     

Novel side chain modified Delta8(14)-15-ketosterols

Synthesis of five novel Delta8(14)-15-ketosterols comprising modified side chains starting from ergosterol is described. Ergosteryl acetate was converted into (22E)-3beta-acetoxy-5alpha-ergosta-8(14),22-dien-15-one through three stages in 32% overall yield; further transformations of the product obtained led to (22E)-3beta-hydroxy-5alpha-ergosta-8(14),22-dien-15-one, (22S,23S)-3beta-hydroxy-22,23-oxido-5alpha-ergost-8(14)-en-15-one, (22R,23R)-3beta-hydroxy-22,23-oxido-5alpha-ergost-8(14)-en-15-one, (22R,23R)-5alpha-ergost-8(14)-en-15-on-3beta,22,23-triol and (22R,23R)-3beta-hydroxy-22,23-isopropylidenedioxy-5alpha-ergost-8(14)-en-15-one. New Delta8(14)-15-ketosterols were evaluated for their cytotoxicity and effects on sterol biosynthesis in human hepatoma Hep G2 cells in comparison with known 3beta-hydroxy-5alpha-cholest-8(14)-en-15-one. Among the compounds tested, (22R,23R)-3beta-hydroxy-22,23-oxido-5alpha-ergost-8(14)-en-15-one was found to be the most potent inhibitor of sterol biosynthesis (IC(50)=0.6+/-0.2microM), whereas (22R,23R)-5alpha-ergost-8(14)-en-15-on-3beta,22,23-triol exhibited the highest cytotoxicity (TC(50)=12+/-3microM at a 24h incubation).     

Metabolism of a novel side chain modified Delta8(14)-15-ketosterol, a potential cholesterol lowering drug: 28-hydroxylation by CYP27A1

The synthetic inhibitors of sterol biosynthesis, 3beta-hydroxy-5alpha-cholest-8(14)-en-15-one and 3beta-hydroxy-24S-methyl-5alpha-cholesta-8(14),22-dien-15-one, are of interest as potential cholesterol lowering drugs. Rapid metabolism of synthetic 15-ketosterols may lead to a decrease, or loss, of their potency to affect lipid metabolism. 3beta-Hydroxy-5alpha-cholest-8(14)-en-15-one is reported to be rapidly side chain oxygenated by rat liver mitochondria. In an attempt to reduce this metabolism, the novel side chain modified 15-ketosterol 3beta-Hydroxy-24S-methyl-5alpha-cholesta-8(14),22-dien-15-one was synthesized. We have examined the metabolism by recombinant human CYP27A1 of this novel side chain modified 3beta-hydroxy-24S-methyl-5alpha-cholesta-8(14),22-dien-15-one and compared the rate of metabolism with that of the previously described 3beta-hydroxy-5alpha-cholest-8(14)-en-15-one. Both sterols were found to be efficiently metabolized by recombinant human CYP27A1. None of the two 15-ketosterols was significantly metabolized by microsomal 7alpha-hydroxylation. Interestingly, CYP27A1-mediated product formation was much lower with the side chain modified 3beta-hydroxy-24S-methyl-5alpha-cholesta-8(14),22-dien-15-one than with the previously described 3beta-hydroxy-5alpha-cholest-8(14)-en-15-one. A surprising finding was that this novel side chain modified sterol was metabolized mainly in the C-28 position by CYP27A1. The data on 28-hydroxylation by human CYP27A1 provide new insights on the catalytic properties and substrate specificity of this enzyme. The finding that 3beta-hydroxy-24S-methyl-5alpha-cholesta-8(14),22-dien-15-one with a modified side chain is metabolized at a dramatically slower rate than the previously described 15-ketosterol with unmodified side chain may be important for future development of synthetic cholesterol lowering sterols.     

New 11(15-->1)abeotaxane, 11(15-->1),11(10-->9)bisabeotaxane and 3,11-cyclotaxanes from Taxus yunnanensis

Chemical examination of the seeds of Chinese yew, Taxus yunnanensis Cheng et L. K. Fu resulted in the isolation of an 11(15-->1)abeotaxane, an 11(15-->1), 11(10-->9)bisabeotaxane and two 3,11-cyclotaxanes. The structures of these new taxoids were established as 13alpha-acetoxy-5alpha-cinnamoyloxy-11(15-->1)abeotaxa-4(20),11-diene-9alpha,10beta,15-triol (1), 20-acetoxy-2alpha-benzoyloxy-4alpha, 5alpha, 7beta, 9alpha, 13alpha-pentahydroxy-11(15-->1), 11(10-->9) bisabeotax-11-eno-10,15-lactone (2), 2alpha,10beta-diacetoxy-5alpha-cinnamoyloxy-9alpha-hydroxy-3,11 -cyclotax-4(20)-en-13-one (3) and 10beta-acetoxy-2alpha,5alpha,9alpha-trihydroxy-3,11-cyclotax-4(20)-en-13-one (4) on the basis of spectral analyses.     

Sesquiterpene constituents from the essential oils of the liverworts Mylia taylorii and Mylia nuda

The essential oils and extracts of Mylia taylorii and M. nuda were investigated by gas chromatography, mass spectrometry, NMR spectroscopy and chemical correlations. Beside several known compounds 13 new constituents including three new carbon skeletons could be identified. Four hydrocarbons with a molecular formula of C₁₅H₂₂ (m/z 202) were identified as myli-4(15)-ene (1), aromadendra-1(10),4(15)-diene (19), aromadendra-4,10(14)-diene (20) and aromadendra-4,9-diene (21). Three oxaspiro-compounds were identified as 7-epi-bourbon-3-en-5,11-oxide (22), guai-3,10(14)-dien-5,11-oxide (23) and guai-3,9-dien-5,11-oxide (24). The absolute configuration of myli-4(15)-en-3-one (5) could be established by chemical correlation. Together with α-taylorione (7) the corresponding 6,11-seco-compound taylopyran (25) with a new carbon skeleton was identified which serves as a precursor to taylocyclane (26) and taylofuran (27). Taynudol (28) contains a new carbon skeleton with a cyclobutenyl structure.     

New polyoxygenated steroids from the Antarctic octocoral Dasystenella acanthina

The chemical study of the Antarctic octocoral Dasystenella acanthina has led to the isolation of the new polyoxygenated steroids (24R,22E)-24-hydroxycholest-4,22-dien-3-one (1), 23-acetoxy-24,25-epoxycholest-4-en-3-one (2), 12beta-acetoxycholest-4-en-3,24-dione (3), 12beta-acetoxy-24,25-epoxycholest-4-en-3-one (4), (22E)-25-hydroxy-24-norcholest-4,22-dien-3-one (5), 3alpha-acetoxy-25-hydroxycholest-4-en-6-one (6), and 3alpha,11alpha-diacetoxy-25-hydroxycholest-4-en-6-one (7), whose structures have been established by spectroscopic analysis. The absolute stereochemistry at C-24 in compound 1 has been determined through the 1H NMR study of the corresponding (R)- and (S)-MPA esters. All the new compounds showed significant activities as growth inhibitors of several human tumor cell lines. In addition, cytostatic and cytotoxic effects were also observed on selected tumor cell lines.     

Antifungal isopimaranes from Hypoestes serpens

Five isopimarane diterpenes (7beta-hydroxyisopimara-8,15-dien-14-one, 14alpha-hydroxyisopimara-7,15-dien-1-one, 1beta,14alpha-dihydroxyisopimara-7,15-diene, 7beta-hydroxyisopimara-8(14),15-dien-1-one and 7beta-acetoxyisopimara-8(14),15-dien-1-one) have been isolated from the leaves of Hypoestes serpens (Acanthaceae). All compounds exhibited antifungal activity against both the plant pathogenic fungus Cladosporium cucumerinum and the yeast Candida albicans; two of them also displayed an acetylcholinesterase inhibition. The structures of the compounds were determined by means of spectrometric methods, including 1D and 2D NMR experiments and MS analysis.     

Diterpenes and sesquiterpenes from the bark of Taxus yunnanensis

Two taxane-type diterpenes, 10beta-acetoxy-2alpha,5alpha,7beta,9alpha-tetrahydroxytaxa-4(20),11-dien-13-one and 2alpha-acetoxy-9alpha-benzoyloxy-5alpha,7beta,10beta,15-tetrahydroxy-11(15-->1)- abeotaxa-4(20),11-dien-13-one, and two new drimane-type sesquiterpenes, 1beta-acetoxy-7-drimen-11alpha-ol-12,11-lactone and 1beta-acetoxy-11,12-epoxy-6-drimen-8alpha,11alpha-diol, were isolated from the bark of Taxus yunnanensis together with 35 known taxane-type diterpenes, a known drimane-type sesquiterpene and a known flavanone.     

Synthesis of (25R)-26-hydroxy-15-ketosterols

(25R)-3beta,26-Dihydroxy-5alpha-cholest-8(14)-en-15-one (1) and (25R)-3beta,26-dihydroxy-5alpha,14beta-cholest-16-en-1 5-one (2) were synthesized from (25R)-3beta,26-dibenzoyloxy-5alpha,14alpha-chole st-16-ene (4). Oxidation of 4 with CrO3-3,5-dimethylpyrazole at -20 degrees C gave (25R)-3beta,26-dibenzoyloxy-5alpha,14alpha-chole st-16-en-15-one (5) along with (25R)-3beta,26-dibenzoyloxy-5alpha-cholest-16alpha+ ++,17alpha-epoxide (6). Oxidation of 5 with selenium dioxide afforded (25R)-3beta,26-dibenzoyloxy-5alpha-cholest-8(14),16-++ +dien-15-one (7) and (25R)-3beta,26-dibenzoyloxy-5alpha,14beta-choles t-16-en-15-one (8). Selective hydrogenation of 7 followed by hydrolysis in alcoholic potassium hydroxide yielded (25R)-3beta,26-dihydroxy-5alpha-cholest-8(14)-en-15-one (1). Hydrolysis of 5 and 8 in alcoholic potassium hydroxide provided (25R)-3beta,26-dihydroxy-5alpha,14beta-cholest-16-en-1 5-one (2).     

Diterpenoids and triterpenoids from Euphorbia guyoniana

Two new compounds with tigliane and cycloartane skeletons: 4,12-dideoxy(4alpha)phorbol-13-hexadecanoate (1) and 24-methylenecycloartane-3,28-diol (2), respectively, in addition of four known diterpenoids and 13 triterpenoids: 3-benzoyloxy-5,15-diacetoxy-9,14-dioxojatropha-6(17),11-diene (4), ent-abieta-8(14),13(15)-dien-16,12-olide (5), ent-8alpha,14alpha-epoxyabieta-11,13(15)-dien-16,12-olide (6), ent-3-hydroxyatis-16(17)-ene-2,14-dione (7), 3beta-hydroxytaraxer-14-en-28-oic acid (8), beta-sitosteryl-3beta-glucopyranoside-6'-O-palmitate (9), multiflorenyl acetate (10), multiflorenyl palmitate (11), peplusol (12), 24-methylenecycloartanol (3), lanosterol (13), euferol (14), butyrospermol (15), cycloartenol (16), obtusifoliol (17), cycloeucalenol (18) and beta-sitosterol (19), were isolated from the roots of Euphorbia guyoniana. Their structures were established on the basis of physical and spectroscopic analysis, including 1D and 2D homo- and heteronuclear NMR experiments (COSY, HSQC, HMBC and NOESY) and by comparison with the literature data.     

Further studies on the inhibition of sterol biosynthesis in animal cells by 15-oxygenated sterols

The chemical syntheses of a number of C27 15-oxygenated sterols and their derivatives have been pursued to permit evaluation of their activity in the inhibition of sterol biosynthesis in animal cells in culture. Described herein are chemical syntheses of 3 alpha-benzoyloxy-5 alpha-cholest-8(14)-en-15-one, 5 alpha-cholest-8(14)-en-3 alpha-ol-15-one, 5 alpha-cholest-8(14)-en-15-one-3 beta-yl pyridinium sulfate, 5 alpha-cholest-8(14)-en-15-one-3 beta-yl potassium sulfate (monohydrate), 5 alpha-cholest-8(14)-en-15-one-3 alpha-yl pyridinium sulfate, 5 alpha-cholest-8(14)-en-3 alpha-yl potassium sulfate (monohydrate), 5 alpha-cholest-8(14)-en3,7,15-trione, 5 alpha-cholest-8(14)-en-15 alpha-ol-3-one, 5 alpha, 14 alpha-cholestan-3 beta, 15 beta-diol diacetate, 5 alpha, 14 beta-cholestan-3 beta, 15 beta-diol diacetate, 5 alpha, 14 alpha-cholestan-3 beta, 15 alpha-diol, 5 alpha, 14 alpha-cholestan-15 alpha-ol-3-one, 5 alpha, 14 beta-cholestan-3 beta, 15 beta-diol, 5 alpha, 14 alpha-cholestan-3,15-dione, and 5 alpha, 14 beta-cholestan-3,5-dione. The effects of 8 of the above compounds and of 5 alpha-cholesta-6,8(14)-dien-3 beta-ol-15-one, 3 beta-he misuccinoyloxy-5 alpha-cholest-8(14)-en-15 one, 3 beta-hexadecanoyloxy-5 alpha-cholest-8(14)-en-15-one, 5 alpha-cholest-8(14)-en-3,15-dione, 5 alpha-cholesta-6,8(14)-dien-3,15-dione, 5 alpha-cholest-8-en-3 beta, 15 alpha-diol, 5 alpha-cholest-7-en-3 beta, 15 alpha-diol, 5 alpha-cholest-8(14)-en-15 alpha-ol-3-one, 5 alpha-cholest-8-en-15 alpha-ol-3-one, and 5 alpha-cholest-7-en-15 alpha-ol-3-one on the synthesis of digitonin-precipitable sterols and on levels of HMG-CoA reductase activity have been investigated and compared with previously published data on 7 other C27 15-oxygenated sterols.     

Cadinane-type sesquiterpenoids from Schisandra bicolor

The phytochemical investigation on the stems of Schisandra bicolor led to the isolation of seven cadinane-type sesquiterpenoids, 2-hydroxy-11,12-dehydrocalamenene (1), ent-T-muurolol (2), (+)-ent-epicubenol (3), (1S,4S)-7,8-dihydroxy-11,12-dehydrocalamenene (4), cadinane-T-cadinol (5), (−)-cadin-4,10(15)-dien-11-oic acid (6), and cadina-4,10(15)-dien-3-one (7). Their structures were determined by extensive analysis of their spectroscopic data. All the isolates were isolated from this species for the first time. The chemotaxonomic significance of these compounds was also summarized.     

Novel P450(17alpha) inhibitors: 17-(2'-oxazolyl)- and 17-(2'-thiazolyl)-androstene derivatives

Twelve 17-(2'-oxazolyl)- and 17-(2'-thiazolyl)-androsta-5,16-diene derivatives were designed and synthesized from 3 beta-acetoxy-pregna-5,16-dien-20-one (1b) as inhibitors of 17 alpha-hydroxylase-C(17,20)-lyase (P450(17 alpha)). Potent inhibitors of this enzyme could be of value as treatment of prostate cancer. Two substituents (methyl and phenyl) were introduced either at their 4'- or 5'-position in order to investigate their structure-activity relationship. Due to the 16,17-double bond, 17-thiazoles were generally obtained in low yield. The pharmacological results showed that the compounds containing 17-(2'-oxazolyl) (14c) and 17-(2'-thiazolyl) (8c) (41.5%) demonstrated reasonable inhibition against P450(17 alpha). Their 3-acetate (13c and 7c) were less potent than their 3-OH counterparts. The introduction of a phenyl or methyl group generally decreased inhibitory activity. Surprisingly, 17-(5'-methyl-2'-thiazolyl) (12a) was the most potent compound in this series and was almost as potent as L-39, which has good antitumor activity.     

The Chemical Constituents of Amoora yunnanensis

Two new sterols, 3β,7α,16β-trihydroxy-stigmast-5,22-diene (1), 3β,7α,16β-trihydroxy-stigmast-5-ene (2), were isolated together with six known compounds, ergosta-5,24(28)-dien-3β,7α-diol (3), ergosta-5,24(28)-dien-3β,7β,16β-triol (4), β-amyrone (5), β-amyrin (6), 11α,12α-epoxy-14-taraxeren-3-one (7), and 6-guaiene-4α,10α-diol (8) from the EtOH extract of the bark of Amoora yunnanensis (H. L. Li) C. Y. Wu. Their structures were deduced on the basis of spectral data.     

Constituents of various wood-rotting basidiomycetes

Phytochemical investigation of n-hexane and methanol extracts of fruiting bodies of the wood-rotting fungi Fomitopsis pinicola. Ganoderma lipsiense, Fomes fomentarius and Gloeophyllum odoratum led to the isolation and identification of several triterpene derivatives and some aromatic compounds derived from lignin. These are the new natural products, namely, pinicolic acid E (16alpha-hydroxy-3-oxolanosta-8,24-dien-21-oic acid) and pinicolol C (3-oxolanosta-7,9(11),24-trien-15alpha,21-diol) from the crust of F. pinicola, ganoderenic acid D [(E)-7beta-hydroxy-3,11,15,23-tetraoxolanosta-8,20(22)-di en-26-oic acid] and ganoderic acid N (7beta,20-dihydroxy-3,11,15,23-tetraoxolanost-8-en-26-oic acid) from G. lipsiense and ergosterol peroxide (5alpha,8alpha-epi-dioxyergost-6-en-3beta-ol) as well as ergost-7-en-3-one from F. fomentarius. From G. odoratum, dehydroeburicoic acid [24-methylene-3-oxolanosta-7,9(11)-dien-21-oic acid], the dimethylacetal of 4,4,14alpha-trimethyl-24-oxo-5alpha-chol-8-en-21-oic acid and some aromatic compounds, of which 1-(4'-methoxyphenyl)-1,2-ethandiol is a new natural product, were isolated. Furthermore, a complete set of 13C NMR data of the steryl esters 3beta-linoleyloxyergosta-7,24(28)-diene, 3beta-linoleyloxyergosta-7,24-diene and 3beta-linoleyloxyergost-7-ene, which could be identified as a mixture in all investigated fungi, could be recorded. It was proved by HPLC and TLC investigations, that the crust on top of the fruiting bodies of F. pinicola consists of lanostane derivatives.     

Sterols from a soft coral, Dendronephthya gigantea as farnesoid X-activated receptor antagonists

Three new steroids 3-oxocholest-1,22-dien-12β-ol (1), 3-oxocholest-1,4-dien-20β-ol (2), 3-oxocholest-1,4-dien-12β-ol (3), and three known steroids (20S)-20-Hydroxyergosta-1,4,24(28)-trien-3-one (4) [7a], 5α,8α-Epidioxycholesta-6,22-dien-3β-ol (5) [10] and 5-cholestene-3β,12β-diol (6) [11] were isolated from a soft coral Dendronephthya gigantea. Their chemical structures and relative stereochemistry were elucidated by the analysis of HRMS and 2-D NMR spectroscopic data. The steroids 1 and 2 showed notable inhibitory activity against farnesoid X-activated receptor (FXR) with IC(50)'s 14 and 15μM.