Discovery of Unprecedented Human Stercobilin Conjugates.

Two unique metabolites (M18 and M19) were detected in feces of human volunteers dosed orally with [14C]inavolisib with a molecular ion of parent plus 304 Da. They were generated in vitro by incubation with fecal homogenates and we have evidence that they are formed chemically and possibly enzymatically. Structural elucidation by high resolution mass spectrometry and nuclear magnetic resonance spectroscopy showed that the imidazole ring of inavolisib was covalently bound to partial structures derived from stercobilin, an end-product of heme catabolism produced by the gut microbiome. The structural difference between the two metabolites was the position of methyl and ethyl groups on the pyrrolidin-2-one moieties. We propose a mechanism of M18 and M19 generation from inavolisib and stercobilin whereby nucleophilic attack from the imidazole ring of inavolisib occurs to the bridging carbon of a stercobilin molecule. The proposed mechanism was supported by computational calculations of molecular orbitals and transition geometry. SIGNIFICANCE STATEMENT: We report the characterization of two previously undescribed conjugates of the phosphoinositide 3-kinase inhibitor inavolisib, generated by reaction with stercobilin, an end-product of heme catabolism produced by the gut microbiome. These conjugates were confirmed by generating them using in vitro fecal homogenate incubation via nonenzymatic and possibly enzymatic reactions. Given the unique nature of the conjugate, it is plausible that it may have been overlooked with other small molecule drugs in prior studies.

Iridoid glucosides in the genus Sutera (Scrophulariaceae) as chemotaxonomic markers in tribe Limoselleae.

From two species of Sutera (S. foetida and S. cordata) (Scrophulariaceae tribe Limoselleae) were isolated three known secoiridoid glucosides (12-14) as well as four iridoid congeners (8-11), all biosynthetically derived from iridodial glucoside (and/or deoxyloganic acid). In addition, two previously unknown compounds were found, namely a terpenoid glucoside lactone (suterolide, 21) and the phenylethanoid glycoside 2''''-O-acetyl-angoroside A (19) as well as verbascoside, echinacoside and tubuloside A(15-17, respectively). Two other species, Jamesbrittenia dissecta and Lyperia antirrhinoides, previously considered to belong to the same genus (Sutera) were shown to be members of two different genera, respectively. Significantly, these two species contained iridoids derived from 8-epi-iridodial (and 8-epideoxyloganic acid), namely aucubin (2), melittoside (3) and acetylharpagide (4). In addition we investigated Melanospermum transvaalense, Lyperia tristis and Microdon dubius likewise from Limoselleae and all of these contained iridoid glucosides from the 8-epi-pathway. Thus, secoiridoid distribution confirms the DNA-based circumscription of Sutera and its sister-group relationship with Manulea. In addition, the results show that the clade including these two genera has a biosynthetic pathway to iridoids fundamentally different from the rest of the tribe and from the whole family Scrophulariaceae.

Occurrence of myo-inositol and alkyl-substituted polysaccharide in the prey-trapping mucilage of Drosera capensis.

The chemical composition of the exudate mucilage droplets of the carnivorous plant Drosera capensis was investigated using nuclear magnetic resonance spectroscopy. The mucilage was found to contain beside a very large molecular weight polysaccharide a significant amount of myo-inositol. It appears that myo-inositol escaped detection due to the commonly applied methodology on the chemical analysis of plant mucilage, such as dialysis, precipitation of polysaccharide component with alcohol, acid hydrolysis and detection of the resultant monosaccharide (aldose) units. The possible functions of myo-inositol in the mucilage droplets and the fate after being washed off from the leaf tentacles are proposed. On the polysaccharide component, the presence of methyl ester and alkyl chain-like moieties could be confirmed. These lipophilic moieties may provide the prey-trapping mucilage with the unique adhesive property onto the hydrophobic insect body parts, as well as onto the nature's well-known superhydrophobic surfaces such as the leaves of the sacred lotus plants. A re-evaluation of the mineral components of the mucilage, reported 40 years ago, is presented from the viewpoints of the current result and plants' natural habitat. A case for re-examination of the well-studied plant mucilaginous materials is made in light of the new findings.

Iridoid glucosides in the genus Veronica (Plantaginaceae) from New Zealand.

Four simple iridoid glucosides, three known esters of catalpol, seven esters of aucubin, and two phenylethanoids were isolated from Veronica hookeri (syn. Hebe ciliolata; Plantaginaceae). Of these, none of four aromatic (p-methoxybenzoyl, isovanilloyl, veratroyl, caffeoyl) 6-O-esters of aucubin and 6″-O-benzoyl mussaenosidic acid, had been reported from nature before. Similarly, three simple iridoid glucosides, two esters of 6-O-rhamnopyranosylcatapol, and two phenylethanoid glucosides, as well as 1-O-benzoyl-3-α-glucuronosylglycerol, and 1-O-ß-benzoyl rutinoside were isolated from Veronica pinguifolia (syn. Hebe pinguifolia). The compound 3″-O-benzoyl-2″-O-caffeoyl 6-O-rhamnopyranosylcatalpol had not been reported previously. The pattern of the structural features of the iridoid glucosides is overlaid onto the latest molecular phylogenetic framework of Veronica sects. Hebe and Labiatoides, and discussed in the context of evolutionary trends.

Protoilludane, Illudane, Illudalane, and Norilludane Sesquiterpenoids from Granulobasidium vellereum.

Two new and seven known sesquiterpene compounds were isolated from an agar plate culture of Granulobasidium vellereum, isolated from a log of Ulmus sp. The two new structures were elucidated with spectroscopic methods as an illudalane derivative, granulolactone (1), and a 15-norilludane, granulodione (9). The acaricidal and insecticidal activities of the isolated compounds were examined in vitro against two major horticultural pests, the two-spotted spider mite Tetranychus urticae and the glasshouse thrips Heliothrips haemorrhoidalis, respectively.

Iridoid glucosides in the endemic Picconia azorica (Oleaceae).

In our continued investigation of plants from the family Oleaceae we have now investigated Picconia azorica endemic to the Azores. Like most species within the family it contains the oleoside-based secoiridoid glucosides ligstroside and oleuropein as the main compounds and in addition verbascoside and echinacoside. As with the previously investigated Picconia excelsa, it also contained the carbocyclic iridoid glucosides involved in the biosynthetic pathway to the oleoside derivatives. However, while P. excelsa contained loganin esterified with some monoterpenoid acids, P. azorica contains similar esters of 7-epi-loganic acid named Picconioside A and B. In addition were found the two 7-O-E/Z-cinnamoyl esters of 7-epi-loganic acid named Picconioside C and D.

Further iridoid glucosides in the genus Manulea (Scrophulariaceae).

From Manulea altissima (Scrophulariaceae) were isolated five known secoiridoid glucosides sweroside, eustomoside, eustoside, secoxyloganin and secologanoside as well as the 4″-O-rhamnopyranosyl-feruloyl ester of adoxosidic acid, named altissimoside. Also, the caffeoyl phenylethanoid glycoside verbascoside was isolated. In addition two previously unknown terpenoid esters of 6ß-hydroxy 8-epi-boschnaloside, named manucoside A and B were isolated from a formerly obtained fraction from the work-up of Manulea corymbosa. The distribution of iridoid glucosides in the Scrophulariaceae is discussed.

Unexpected secoiridoid glucosides from Manulea corymbosa.

From an extract of Manulea corymbosa were isolated four known secoiridoid glucosides (1-4), 10 new monoterpenoid esters of secologanol, namely, manuleosides A-I (5-11, 13, and 14) and dimethyl rhodanthoside A (12), and four new phenylpropanoid esters of carbocyclic iridoid glucosides, manucorymbosides I-IV (15-18). Also, the caffeoyl phenylethanoid glycoside verbascoside was isolated. The presence of secoiridoids apparently derived from loganic acid in the family Scrophulariaceae is unprecedented and greatly unexpected.

Direct and unbiased information recovery from liquid chromatography-mass spectrometry raw data for phenotype-differentiating metabolites based on screening window coefficient of ion currents.

A reworking of a data mining strategy, in which statistical treatment of raw data from liquid chromatography-mass spectrometry (LC-MS) precedes recognition of chromatographic peaks, is presented. In this algorithm the tR-m/z plane of LC-MS data is divided into equal-sized segments of twelve seconds by one m/z unit each, and the total ion currents in corresponding segments as specified by the tR-m/z pair from multiple LC-MS runs are evaluated to generate mean ion currents (µ) and standard deviations (σ). The µ's and σ's of the segments, derived from contrasting classes of LC-MS data set (e.g., resistant-susceptible, case-control, etc.), are used to calculate the Z-factor (screening window coefficient) which is in turn used to rank the segments. Chromatographic peaks are recognized only where the ion currents are shown to differentiate the classes. The result-reporting format enables detection of positive as well as negative correlations between ion intensities and biological traits under study and thus points to the presence of potentially phenotype-discriminating metabolites. Examples of data analyses are presented, in which ions that may distinguish resistant and susceptible species of Aesculus to the leaf-miner Cameraria ohridella were detected.

Amides and an alkaloid from Portulaca oleracea.

A total of 16 phenolic compounds, including one new and five known N-cinnamoyl phenylethylamides, one new pyrrole alkaloid named portulacaldehyde, five phenylpropanoid acids and amides, and derivatives of benzaldehyde and benzoic acid, were isolated and identified from a polar fraction of an extract of Portulaca oleracea. Their structures were determined through spectroscopic analyses.

Iridoid and phenylethanoid glycosides in the New Zealand sun hebes (Veronica; Plantaginaceae).

The sun hebes are a small clade of New Zealand Veronica formerly classified as Heliohebe. The water-soluble compounds of Veronica pentasepala, Veronica raoulii and Veronica hulkeana were studied and 30 compounds including 15 iridoid glucosides, 12 phenylethanoid glycosides, the acetophenone glucoside pungenin, the mannitol ester hebitol II and mannitol were isolated. Of these, five were previously unknown in the literature: dihydroverminoside and 3,3',4,4'-tetrahydroxy-α-truxillic acid 6-O-catalpyl diester, named heliosepaloside, as well as three phenylethanoid glycoside esters heliosides D, E and F, all derivatives of aragoside. The esters of cinnamic acid derivatives with iridoid and phenylethanoid glycosides and an unusually high concentration of verminoside were found to be the most distinctive chemotaxonomic characters of the sun hebes. The chemical profiles of the species were compared and used to assess the phylogenetic relationships in the group.

Iridoid and phenylethanoid glucosides from Veronica lavaudiana.

From an extract of Veronica (sect. Hebe) lavaudiana we have identified mannitol and isolated 11 iridoid glucosides, the carbohydrate ester hebitol II, and four phenylethanoid glycoside esters. Five of the iridoid glycosides are new; of these, lavaudiosides A, B, and C (2a, 3a, and 4) are 1-mannityl esters of 8-epiloganic acid, while 7e and 7f are 6'-O-caffeoyl derivatives of catalpol. The new phenylethanoid glycoside esters, heliosides A, B, and C (8b-d), are 6'-xylosyl derivatives of aragoside. The structures of the new compounds were elucidated mainly by spectroscopic analysis, but also by chemical degradation. We also demonstrated that the structures of the known glycosides globularitol and hebitols I and II should be revised. These compounds are derivatives of mannitol and not glucitol as previously believed.

Phenylethanoid and iridoid glycosides in the New Zealand snow hebes (Veronica, Plantaginaceae).

Snow hebes are the alpine cushion-forming plants of New Zealand Veronica, formerly classified as Chionohebe. The chemical compositions of Veronica pulvinaris and Veronica thomsonii were studied and 33 water-soluble compounds were isolated. The structures of 14 previously unknown esters of phenylethanoid glycosides were elucidated by spectroscopic analyses. Further, eight known phenylethanoids, nine iridoids, 6'-feruloyl-sucrose and mannitol are also reported. It was found that the iridoid profile of the snow hebes was different from the other species of Veronica in New Zealand but similar to the alpine Northern Hemisphere representatives of the genus.

Antibacterial and antifungal activity of cicerfuran and related 2-arylbenzofurans and stilbenes.

Cicerfuran, 2-(2-methoxy-4,5-methylenedioxyphenyl)-6-hydroxybenzofuran, is an antifungal phytoalexin previously isolated from the roots of chickpea, Cicer spp. The synthesis of cicerfuran, five 2-arylbenzofuran analogues and nine stilbene intermediates was reported recently. The antimicrobial activities of these compounds were evaluated against two species of bacteria, Bacillus subtilis and Pseudomonas syringae, and four species of filamentous fungi, Aspergillus niger, Botrytis cinerea, Cladosporium herbarum and Monilinia aucupariae. Stilbenes with a free hydroxyl group were active against both bacteria and fungi with MICs in the range 25-100microg/ml. Cicerfuran was the only 2-arylbenzofuran that showed antimicrobial activity with MICs as low as 25microg/ml. Some aspects of the structure-activity relationship are discussed.

Chemical constituents and antimicrobial activity of medicinal plants from Ghana: Cassia sieberiana, Haematostaphis barteri, Mitragyna inermis and Pseudocedrela kotschyi.

The antimicrobial activity of the sequential n-hexane, acetone and 50% aqueous methanol extracts of leaves, stem bark and roots of four species of medicinal plants, Cassia sieberiana DC. (Leguminosae), Haematostaphis barteri Hook. f. (Anacardiaceae), Mitragyna inermis (Willd.) O. Kuntze (Rubiaceae) and Pseudocedrela kotschyi (Schweinf.) Harms (Meliaceae), from Ghana were tested against Bacillus subtilis, Pseudomonas syringae and Cladosporium herbarum using TLC direct-autobiographic methods. Extracts from leaves, stem bark and roots of the four species gave a positive result against at least one test organism. Twelve of the 36 extracts were active against B. subtilis, four extracts were active against P. syringae and six were active against C. herbarum. Preliminary chemical analysis revealed the presence of flavonoids, stilbenes and alkaloids. This is the first report of a stilbene from the Anacardiaceae.

Benzaldehyde derivatives from Sarcodontia crocea.

Through bioactivity-guided fractionation, two volatile benzaldehyde derivatives ( 1 and 2) were isolated from a cellulose-matrix culture of the Basidiomycete Sarcodontia crosea (syn. S. setosa), where the fungus was grown on a growth medium-impregnated thick chromatographic paper. The structures of 1 and 2 were elucidated mainly by 2D NMR techniques as 4-(furan-3-yl)benzaldehyde and 4-(5-oxotetrahydrofuran-3-yl)benzaldehyde, respectively. Compound 1 is a new natural product, while 2 has been synthesized previously. They are considered to contribute to the odor of the fungus, which is an important character for identification of the species. They were weakly antifungal toward several phytopathogenic fungi but less potent against bacteria in microdilution assays.

Inhibitory activities of lichen-derived compounds against methicillin- and multidrug-resistant Staphylococcus aureus.

The inhibitory effects of selected phenolic lichen substances were tested against a panel of methicillin- and multidrug-resistant Staphylococcus aureus. Depsidones with long alkyl chains on both of the aromatic rings were consistently active against the strains tested, comparable to or better than the level of clinically used antibacterial drugs. A similar level of activity was also observed for rhizocarpic acid. The previously described cytotoxic pentacyclic compound hybocarpone was by far the most active, exhibiting minimum inhibitory concentrations (MICs) of 4-8 microg/mL (8.13-16.3 microM) against a range of multidrug efflux pump expressing strains of S. aureus.

Phenylethanoids, iridoids and a spirostanol saponin from Veronica turrilliana.

From the aerial parts of Veronica turrilliana two phenylethanoid glycosides, turrilliosides A and B and a steroidal saponin, turrillianoside were isolated and their structures elucidated as beta-(3,4-dihydroxyphenyl)ethyl-4-O-E-caffeoyl-O-[beta-glucopyranosyl-(1-->4)-alpha-rhamnopyranosyl-(1-->6)]-beta-glucopyranoside, beta-(3,4-dihydroxyphenyl)ethyl-4-O-E-caffeoyl-[6-O-E-feruloyl-beta-glucopyranosyl-(1-->4)-alpha-rhamnopyranosyl-(1-->6)]-beta-glucopyranoside and (23S,25S)-12beta,23-dihydroxyspirost-5-en-3beta-yl O-alpha-rhamnopyranosyl-(1-->4)-beta-glucopyranoside, respectively. Furthermore, eight known glucosides are reported namely, catalpol, catalposide, verproside, amphicoside, isovanilloylcatalpol, aucubin, arbutin, and 6-O-E-caffeoylarbutin, the latter two for the first time in the genus Veronica. The two phenylethanoid glycosides were found to be potent DPPH radical scavengers. All of the tested compounds were inactive against the representative species of fungi and bacteria.

Serialynic acid, a new phenol with an isopentenyne side chain from Antrodia serialis.

A new phenolic compound serialynic acid was isolated from an agar culture of the basidiomycete Antrodia serialis, through bioactivity-guided fractionations. It showed weak growth inhibitory activity towards phytopathogenic fungi and a dose-independent anti-Pythium graminicola activity.

Insect-antifeedant and antibacterial activity of diterpenoids from species of Plectranthus.

Bio-assay guided fractionation of an acetone extract of leaf material from Plectranthus saccatus Benth. resulted in the isolation of a beyerane diterpenoid. This compound, characterised by spectroscopic methods as ent-3beta-(3-methyl-2-butenoyl)oxy-15-beyeren-19-oic acid, showed insect antifeedant activity against Spodoptera littoralis. Known quinonoid abietane diterpenoids obtained from new sources included a mixture of the (4R,19R) and (4R,19S) diastereoisomers of coleon A from P. aff. puberulentus J.K. Morton, coleon A lactone from P. puberulentus J.K. Morton, and coleon U and coleon U quinone from P. forsteri 'Marginatus' Benth. These compounds, and the crude acetone extracts from the leaf surfaces of 11 species of Plectranthus, were tested for antifeedant activity against S. littoralis, antibacterial activity against Bacillus subtilis and Pseudomonas syringae and antifungal activity against Cladosporium herbarum. The coleon A mixture showed potent antifeedant activity against S. littoralis, whereas coleon U showed the greatest antimicrobial activity.