Inhibition of multidrug-resistant MCF-7 breast cancer cells with combinations of clinical drugs and resin glycosides from Operculina hamiltonii.

The jalap roots, Operculina hamiltonii D.F. Austin & Staples (Convolvulaceae), are extensively commercialized as a depurative and laxative remedy in traditional medicine of the north and northeast regions of Brazil. The purification by recycling HPLC and structure elucidation of three new acyl sugars or resin glycosides are described here from a commercial product made of powdered roots. Three macrocyclic structures of a tetrasaccharide of (11S)-hydroxyhexadecanoic acid, operculinic acid C (1), the undescribed hamiltonins II and III (3 and 4), in addition to the known batatinoside III (5), presented a diastereoisomeric relationship as one residue of n-dodecanoic acid esterified the oligosaccharide core on a different position in each compound. Furthermore, hamiltonin IV (6) was characterized as an ester-type homodimer of acylated operculinic acid C with the same substitution pattern identified in hamiltonins II (3) and III (4) for each of the dimer subunits. All the isolated resin glycosides did not display any intrinsic cytotoxicity (IC50 > 25 µM). However, a combination of the individual isolated compounds 3-6 (1-50 µM) demonstrated an enhancement of cytotoxic effects with sublethal doses of vinblastine and podophyllotoxin (0.003 µM) in multidrug-resistant breast carcinoma epithelial cells (MCF-7/Vin).

Hederifolic acids A-D, hepta and hexasaccharides from the resin glycosides of Ipomoea hederifolia.

Scarlet morning glory, Ipomoea hederifolia L. (Convolvulaceae), is an ornamental vine native to the Americas with oxytocic, cytotoxic, antipsychotic, anti-inflammatory, antioxidant, and antimicrobial properties. A chemical study of the glycosidic acids from the resin glycosides contained in the aerial parts was carried out, through their isolation as peracetylated derivatives, by recycling preparative liquid chromatography. Structure elucidation was performed by HR-MS in accordance with NMR. Four peracetylated derivatives of glycosidic acids, named hederifolic acids A-D, were identified as heptaglycosides and hexaglycosides linked to 3S,12S-dihydroxyheptadecanoic acid or 12 S-hydroxyheptadecanoic acid. Consequently, hederifolic acids B and D were found to be dehydroxylated homologs at C-3 of the fatty acid aglycones of hederifolic acids A and C, respectively.

Resin Glycosides from Operculina hamiltonii and Their Synergism with Vinblastine in Cancer Cells.

Operculina hamiltonii is a vine native to the north and northeast region of Brazil, where its roots are traded as a depurative and laxative remedy with the name of Brazilian jalap in traditional medicine. Procedures for the isolation, purification by recycling HPLC, and structure elucidation of three undescribed resin glycosides are presented herein. Hamiltonin I (1) represents a macrocyclic structure of a tetrasaccharide of (11S)-hydroxyhexadecanoic acid. Additionally, two acyclic pentasaccharides, named hamiltoniosides I (2) and II (3), were also isolated, which are related structurally to the known compounds 4 and 5, macrocyclic lactone-type batatinosides. The tetrasaccharide core of 1 was diacylated by n-decanoic acid and the unusual n-hexadecanoic acid moiety, while the pentasaccharides 2-5 were esterified by one unit of n-decanoic or n-dodecanoic acid. All the isolated compounds were found to be inactive as cytotoxic agents. However, when they were evaluated (1-25 µM) in combination with a sublethal concentration of the anticancer agent vinblastine (0.003 µM), a significant enhancement of the resultant cytotoxicity was produced, especially for multidrug-resistant breast carcinoma epithelial cells. Such combined synergistic potency may be beneficial for chemotherapy, making resin glycosides potential candidates for drug repurposing of conventional chemotherapeutic drugs to reduce their side effects.

Distribution of 5,6-dihydro-α-pyrones by electrospray ionization ion trap mass spectrometry in different aerial parts of Hyptis monticola.

Hyptis monticola Mart. ex Benth. (Lamiaceae) is an endemic species of altitude regions of Brazil. From the leaves of this plant, two 5,6-dihydro-α-pyrones, named monticolides A and B, have been reported as cytotoxic agents against different tumor cell lines. The isolation by high-speed countercurrent chromatography in combination with recycling preparative high-performance liquid chromatography of the undescribed monticolides C-F is presented. These compounds corresponded to a series of related monticolide derivatives differing from each other by the number of acyl substituents. Their characterization by mass spectrometry and nuclear magnetic resonance is also presented, in conjunction with an evidence by a simple chemical correlation for their absolute stereochemistry. The distribution of these chemical markers in extracts of flowers, leaves and branches collected in different seasons by electrospray ionization ion trap mass spectrometry in positive mode was analyzed. Multivariate data analyses indicated that seasonality affects monticolide concentrations in different organs of the aerial parts. Monticolides A-F seem to be present as the original markers of the analyzed plant. However, mono-, di- and triacetylated monticolides can undergo acid-catalyzed transesterifications and their natural yields estimated were affected during the isolation procedures.

Dihydro-furanones from Hyptis species: Chemical correlations and DFT-NMR/ECD calculations for stereochemical assignments.

Dihydro-furanones are bioactive compounds isolated from various plants, marine fungi, and sponges. The present investigation describes the isolation by recycling HPLC and structural characterization by NMR of four previously undescribed 2(5H)-furanones, monticofuranolide A and pectinolides N-P, one phenylpropanoid, rosmarinic acid, and five known flavonoids, in addition to the undescribed natural flavonoid, 2R,3R-dihydrogossipetin or 5,7,8,3',4'-pentahydroxy flavanonol, from collections of H. monticola Mart. ex Benth and Hyptis pectinata (L.) Poit. Chemical correlations, resembling the biogenetic relationship of the isolated 2(5H)-furanones with their 5,6-dihydro-2H-pyran-2-one precursors, were accomplished to confirm their absolute configuration. Density functional theory-NMR/ECD calculations have been used to solve the absolute configuration for this type of compounds.

Dereplication of podophyllotoxin and related cytotoxic lignans in Hyptis verticillata by ultra-high-performance liquid chromatography tandem mass spectrometry.

INTRODUCTION: Hyptis verticillata Jacq. (Lamiaceae) is a Mexican medicinal plant for the treatment of skin infections and illness affecting the respiratory and gastrointestinal systems. OBJECTIVE: To associate the efficient resolution provided by ultra-high-performance liquid chromatography combined to the accuracy of a hybrid Fourier-transform (FT) mass spectrometer in order to dereplicate podophyllotoxin-type lignans in a plant extract. METHODS: An ultra-high-performance liquid chromatography-photodiode array-high resolution electrospray ionisation tandem mass spectrometry (UHPLC-PDA-HRESI-MS/MS) method was applied in an Orbitrap hybrid FT spectrometer for dereplication of podophyllotoxin and related cytotoxic lignans in wild bushmint. This procedure included high-resolution mass values for positively charged ions [M + H]+ and [M + NH4 ]+ , MS/MS data, and comparison of UV maxima and retention times with pure compounds. RESULTS: Podophyllotoxin in addition to seven aryltetralins, four arylnaphthalenes, and one dibenzylbutyrolactone were dereplicated from the methanol extract in a short-time analysis (5 min). 4'-O-Demethyl-dehydro-deoxypodophyllotoxin was identified as a new natural product. CONCLUSION: The applied UHPLC-MS/MS dereplication method is suitable for a rapid analysis of podophyllotoxin-type lignans and the resulting chemical fingerprinting could be valuable in quality control of herbal drugs and their phytopharmaceuticals.

Resin Glycosides from the Roots of Operculina macrocarpa (Brazilian Jalap) with Purgative Activity.

Analysis of the methanol-soluble resin glycosides from the roots of Operculina macrocarpa was assessed by generating NMR profiles of five glycosidic acids obtained through saponification, acetylation, and recycling HPLC purification. Operculinic acid H (1), two novel hexasaccharides, operculinic acids I (2) and J (3), the known purgic acid A (4), and a quinovopyranoside of (-)-(7 R)-hydroxydecanoic acid, operculinic acid K (5), were isolated. Three intact resin glycosides related to 1, the novel macrocarposidic acids A (6) and B (7), in addition to the previously known macrocarposidic acid C (8), were also purified with isovaleroyl, tigloyl, and exogonoyl [(3 S,9 R)-3,6:6,9-diepoxydecanoyl] groups as esterifying residues. A selective intramolecular lactonization was produced to generate a macrocyclic artifact (17) during acetylation of 1, resembling the distinctive structure of the Convolvulaceous resin glycosides.

Antifungal Phenylpropanoid Glycosides from Lippia rubella.

Lippia species share various pharmacological activities and are used in traditional cooking and medicine worldwide. Combined chromatographic techniques such as column chromatography, high-performance liquid chromatography, and countercurrent chromatography led to the purification of two new antifungal phenylpropanoid glycosides, lippiarubelloside A (1) and lippiarubelloside B (2), by bioactivity-directed fractionation of an ethanol-soluble extract from Lippia rubella, in addition to the known active related compounds forsythoside A (3), verbascoside (4), isoverbascoside (5), and poliumoside (6). The structures of compounds 1 and 2 were determined by comparison of their NMR spectroscopic data with the prototype active compound 4. Cryptococcus neoformans, which causes opportunistic lung infections, was sensitive to compounds 1-6 in the concentration range of 15-125 µg/mL. A synergistic effect (FICindex = 0.5) between 3 and amphotericin B was demonstrated. The glycosylated flavonoids pectolinarin (7), linarin (8), and siparunoside (9) were also isolated.

Structure Elucidation, Conformation, and Configuration of Cytotoxic 6-Heptyl-5,6-dihydro-2 H-pyran-2-ones from Hyptis Species and Their Molecular Docking to α-Tubulin.

Cytotoxic 6-heptyl-5,6-dihydro-2 H-pyran-2-ones are chemical markers of Hyptis (Lamiaceae) and are responsible for some of the therapeutic properties of species with relevance to traditional medicine. The present investigation describes the isolation of known pectinolides A-C (1-3), in addition to the new pectinolides I-M (4-8), from two Mexican collections of H. pectinata by HPLC. The novel biosynthetically related monticolides A (9) and B (10) were also isolated by high-speed countercurrent chromatography from H. monticola, an endemic species of the Brazilian southeastern high-altitude regions. A combination of chemical correlations, chiroptical measurements, and Mosher ester NMR analysis was used to confirm their absolute configuration. The utility of DFT-NMR chemical shifts and JH-H calculations was assessed for epimer differentiation. Molecular docking studies indicated that 6-heptyl-5,6-dihydro-2 H-pyran-2-ones have a high affinity for the pironetin-binding site of α-tubulin, which may be a possible mechanism contributing to the cytotoxic potential of these small and flexible molecules.

Morning glory resin glycosides as α-glucosidase inhibitors: In vitro and in silico analysis.

Twenty-seven individual resin glycosides from the morning glory family (Convolvulaceae) were evaluated for their α-glucosidase inhibitory potential. Four of these compounds displayed an inhibitory activity comparable to acarbose, which was used as a positive control. Molecular modeling studies performed by docking analysis were accomplished to predict that the active compounds and acarbose bind to the α-1,4-glucosidase enzyme catalytic site of MAL12 from the yeast Saccharomyces cerevisiae through stable hydrogen bonds primarily with the amino acid residues HIS279 and GLN322. Docking studies with the human maltase-glucoamylase (MGAM) also identified binding modes for resin glycosides inside the catalytic site in the proximity of TYR1251. These results postulate that resin glycosides may be a source of phytotherapeutic agents with antihyperglycemic properties for the prophylaxis and treatment of non-insulin-dependent type 2 diabetes mellitus.

HPLC-MS profiling of the multidrug-resistance modifying resin glycoside content of Ipomoea alba seeds

ABSTRACT High performance liquid chromatography profiling with mass spectrometry detection was applicable to identify known and novel multidrug-resistance glycolipid inhibitors from the complex resin glycosides mixture of Ipomoea alba L., Convolvulaceae, seeds. Albinosides X and XI were purified by recycling liquid chromatography and their structural elucidation was accomplished by nuclear magnetic resonance. Albinoside XI exerted a strong potentiation of vinblastine susceptibility in multidrug-resistant human breast carcinoma cells.

Complementarity of DFT Calculations, NMR Anisotropy, and ECD for the Configurational Analysis of Brevipolides K-O from Hyptis brevipes.

Brevipolides K-O (1-5), five new cytotoxic 6-(6'-cinnamoyloxy-2',5'-epoxy-1'-hydroxyheptyl)-5,6-dihydro-2H-pyran-2-ones (IC50 values against six cancer cell lines, 1.7-10 µM), were purified by recycling HPLC from Hyptis brevipes. The structures, containing a distinctive tetrahydrofuran ring, were established by comprehensive quantum mechanical calculations and experimental spectroscopic analysis of their NMR and ECD data. Detailed analysis of the experimental NMR 1H-1H vicinal coupling constants in comparison with the corresponding DFT-calculated values at the B3LYP/DGDZVP level confirmed the absolute configuration of 3 and revealed its conformational preferences, which were further strengthened by NOESY correlations. NMR anisotropy experiments by the application of Mosher's ester methodology and chemical correlations were also used to conclude that this novel brevipolide series (1-5) share the same absolute configuration corresponding to C-6(R), C-1'(S), C-2'(R), C-5'(S), and C-6'(S).

Resin Glycosides from Ipomoea alba Seeds as Potential Chemosensitizers in Breast Carcinoma Cells.

Multidrug resistance is the expression of one or more efflux pumps, such as P-glycoprotein, and is a major obstacle in cancer therapy. The use of new potent and noncytotoxic efflux pump modulators, coadministered with antineoplastic agents, is an alternative approach for increasing the success rate of therapy regimes with different drug combinations. This report describes the isolation and structure elucidation of six new resin glycosides from moon vine seeds (Ipomoea alba) as potential mammalian multidrug-resistance-modifying agents. Albinosides IV-IX (1-6), along with the known albinosides I-III (7-9), were purified from the CHCl3-soluble extract. Degradative chemical reactions in combination with NMR spectroscopy and mass spectrometry were used for their structural elucidation. Four new glycosidic acids, albinosinic acids D-G (10-13), were released by saponification of natural products 3-6. They were characterized as tetrasaccharides of either convolvulinolic (11S-hydroxytetradecanoic) or jalapinolic (11S-hydroxyhexadecanoic) acids. The potentiation of vinblastine susceptibility in multidrug-resistant human breast carcinoma cells of albinosides 1-6 was evaluated by modulation assays. The noncytotoxic albinosides VII (4) and VIII (5), at a concentration of 25 µg/mL, exerted the strongest potentiation of vinblastine susceptibility, with a reversal factor (RFMCF-7/Vin+) of 201- and >2517-fold, respectively.

Resin glycosides from Ipomoea wolcottiana as modulators of the multidrug resistance phenotype in vitro.

Recycling liquid chromatography was used for the isolation and purification of resin glycosides from the CHCl3-soluble extracts prepared using flowers of Ipomoea wolcottiana Rose var. wolcottiana. Bioassay-guided fractionation, using modulation of both antibiotic activity against multidrug-resistant strains of Gram-negative bacteria and vinblastine susceptibility in breast carcinoma cells, was used to isolate the active glycolipids as modulators of the multidrug resistance phenotype. An ester-type dimer, wolcottine I, one tetra- and three pentasaccharides, wolcottinosides I-IV, in addition to the known intrapilosin VII, were characterized by NMR spectroscopy and mass spectrometry. In vitro assays established that none of these metabolites displayed antibacterial activity (MIC>512 µg/mL) against multidrug-resistant strains of Escherichia coli, and two nosocomial pathogens: Salmonella enterica serovar Typhi and Shigella flexneri; however, when tested (25 µg/mL) in combination with tetracycline, kanamycin or chloramphenicol, they exerted a potentiation effect of the antibiotic susceptibility up to eightfold (64 µg/mL from 512 µg/mL). It was also determined that these non-cytotoxic (CI50>8.68 µM) agents modulated vinblastine susceptibility at 25 µg/mL in MFC-7/Vin(+) cells with a reversal factor (RFMCF-7/Vin(+)) of 2-130 fold.

Bufadienolides from parotoid gland secretions of Cuban toad Peltophryne fustiger (Bufonidae): Inhibition of human kidney Na(+)/K(+)-ATPase activity.

Parotoid gland secretions of toad species are a vast reservoir of bioactive molecules with a wide range of biological properties. Herein, for the first time, it is described the isolation by preparative reversed-phase HPLC and the structure elucidation by NMR spectroscopy and/or mass spectrometry of nine major bufadienolides from parotoid gland secretions of the Cuban endemic toad Peltophryne fustiger: ψ-bufarenogin, gamabufotalin, bufarenogin, arenobufagin, 3-(N-suberoylargininyl) marinobufagin, bufotalinin, telocinobufagin, marinobufagin and bufalin. In addition, the secretion was analyzed by UPLC-MS/MS which also allowed the identification of azelayl arginine. The effect of arenobufagin, bufalin and ψ-bufarenogin on Na(+)/K(+)-ATPase activity in a human kidney preparation was evaluated. These bufadienolides fully inhibited the Na(+)/K(+)-ATPase in a concentration-dependent manner, although arenobufagin (IC50 = 28.3 nM) and bufalin (IC50 = 28.7 nM) were 100 times more potent than ψ-bufarenogin (IC50 = 3020 nM). These results provided evidence about the importance of the hydroxylation at position C-14 in the bufadienolide skeleton for the inhibitory activity on the Na(+)/K(+)-ATPase.

DFT 1H-1H coupling constants in the conformational analysis and stereoisomeric differentiation of 6-heptenyl-2H-pyran-2-ones: configurational reassignment of synargentolide A.

Density functional theory (DFT) (1) H-(1) H NMR coupling constant calculations, including solvation parameters with the polarizable continuum model B3LYP/DGDZVP basis set together with the experimental values measured by spectral simulation, were used to predict the configuration of hydroxylated 6-heptenyl-5,6-dihydro-2H-pyran-2-ones 1, 2, 4, and 7, allowing epimer differentiation. Modeling of these flexible compounds requires the inclusion of solvation models that account for stabilizing interactions derived from intramolecular and intermolecular hydrogen bonds, in contrast with peracetylated derivatives (3, 5, and 6) in which the solvation consideration can be omitted. Using this DFT NMR integrated approach as well as spectral simulation, the configurational reassignment of synargentolide A (8) was accomplished by calculations in the gas phase among four possible diastereoisomers (8-11). Calculated (3) JH,H values established its configuration as 6R-[4'S,5'S,6'S-(triacetyloxy)-2E-heptenyl]-5,6-dihydro-2H-pyran-2-one (8), in contrast with the incorrect 6R,4'R,5'R,6'R-diastereoisomer previously proposed by synthesis (12). Application of this approach increases the probability for successful enantiospecific total syntheses of flexible compounds with multiple chiral centers.

Jalapinoside, a macrocyclic bisdesmoside from the resin glycosides of Ipomea purga, as a modulator of multidrug resistance in human cancer cells.

The first macrocyclic bisdesmoside resin glycoside, jalapinoside (4), was purified by preparative-scale recycling HPLC from the MeOH-soluble extracts of Ipomoea purga roots, the officinal jalap. Purgic acid C (3), a new glycosidic acid of ipurolic acid, was identified as 3-O-ß-d-quinovopyranoside, 11-O-ß-d-quinovopyranosyl-(1→2)-O-ß-d-glucopyranosyl-(1→3)-O-[ß-d-fucopyranosyl-(1→4)]-O-α-l-rhamnopyranosyl-(1→2)-O-ß-d-glucopyranosyl-(1→2)-O-ß-d-quinovopyranoside (3S,11S)-dihydroxytetradecanoic acid. The acylating residues of this core were acetic, (+)-(2S)-methylbutanoic, and dodecanoic acids. The site of lactonization was defined as C-3 of the second saccharide moiety. Reversal of multidrug resistance by this noncytotoxic compound was evaluated in vinblastine-resistant human breast carcinoma cells.

Batatins VIII-XI, glycolipid ester-type dimers from Ipomoea batatas.

Sweet potato (Ipomoea batatas) is native to the tropics of Central and South America, where many varieties have been consumed for more that 5000 years. In developing countries, this crop is a recognized effective food for fighting malnutrition. Purification of the minor lipophilic glicolipids found in the n-hexane-soluble resin glycosides from the white-skinned variety was performed by preparative-scale recycling HPLC. Application of column overload, peak shaving, heart cutting, and recycling techniques permitted the purification of four new oligosaccharide ester-type dimer derivatives of jalapinolic acid, batatins VIII-XI (1-4). The structural characterization of these complex lipo-oligosaccharides was performed through NMR spectroscopy and MS, indicating that batatins VIII-XI (1-4) possess an oligomeric structure consisting of two pentasaccharide units of the known simonic acid B.