Nematicidal Activity of 20-Deoxyingenol-3-angelate from Euphorbia peplus Latex Through Protein Kinase C Isotype TPA-1.

The latex of Euphorbia peplus and its major component 20-deoxyingenol-3-angelate (DI3A) displayed significant nematicidal activity against Caenorhabditis elegans and Panagrellus redivivus. DI3A treatment inhibited the growth and development of nematodes and caused significantly negative effects on locomotion behavior, reproduction, and accumulation of reactive oxygen species. Transcriptome analysis indicated that differential expression genes in DI3A-treated C. elegans were mainly associated with the metabolism, growth, and development process, which were further confirmed by RT-qPCR experiments. The expression level of TPA-1 gene encoding a protein kinase C isotype was obviously upregulated by DI3A treatment, and knockdown of TPA-1 by RNAi technology in the nematode could relieve the growth-inhibitory effect of DI3A. Metabolic analysis indicated that DI3A was hardly metabolized by C. elegans, but a glycosylated indole derivative was specifically accumulated likely due to the activation of detoxification. Overall, our findings suggested that DI3A from E. peplus latex exerted a potent nematicidal effect through the gene TPA-1, which provides a potential target for the control of nematodes and also suggests the potential application value of E. peplus latex and DI3A as botanical nematicides.

Chemical Transformation of B- to A-type Proanthocyanidins and 3D Structural Implications.

In nature, proanthocyanidins (PACs) with A-type linkages are relatively rare, likely due to biosynthetic constraints in the formation of additional ether bonds to be introduced into the more common B-type precursors. However, A-type linkages confer greater structural rigidity on PACs than do B-type linkages. Prior investigations into the structure-activity relationships (SAR) describing how plant-derived PACs with B- and complex AB-type linkages affect their capacity for dentin biomodification indicate that a higher ratio of double linkages leads to a greater interaction with dentin type I collagen. Thus, A-type PACs emerge as particularly intriguing candidates for interventional functional biomaterials. This study employed a free-radical-mediated oxidation using DPPH to transform trimeric and tetrameric B-type PACs, 2 and 4, respectively, into their exclusively A-type linked analogues, 3 and 5, respectively. The structures and absolute configurations of the semisynthetic products, including the new all-A-type tetramer 5, were determined by comprehensive spectroscopic analysis. Additionally, molecular modeling investigated the conformational characteristics of all trimers and tetramers, 1-5. Our findings suggest that the specific interflavan linkages significantly impact the flexibility and low-energy conformations of the connected monomeric units, which conversely can affect the bioactive conformations relevant for dentin biomodification.

Modulatory role of terminal monomeric flavan-3-ol units in the viscoelasticity of dentin.

Flavan-3-ol monomers are the building blocks of proanthocyanidins (PACs), natural compounds from plants shown to mediate specific biologic activities on dentin. While the stereochemistry of the terminal flavan-3-ols, catechin (C) versus epicatechin (EC), impacts the biomechanical properties of the dentin matrix treated with oligomeric PACs, structure-activity relationships driving this bioactivity remain elusive. To gain insights into the modulatory role of the terminal monomers, two highly congruent trimeric PACs from Pinus massoniana only differing in the stereochemistry of the terminal unit (Trimer-C vs. Trimer-EC) were prepared to evaluate their chemical characteristics as well as their effects on the viscoelasticity and biostability of biomodified dentin matrices via infrared spectroscopy and multi-scale dynamic mechanical analyses. The subtle alteration of C versus EC as terminal monomers lead to distinct immediate PAC-trimer biomodulation of the dentin matrix. Nano- and micro-dynamic mechanical analyses revealed that Trimer-EC increased the complex moduli (0.51 GPa) of dentin matrix more strongly than Trimer-C (0.26 GPa) at the nanoscale length (p < 0.001), whereas the reverse was found at the microscale length (p < .001). The damping capacity (tan δ) of dentin matrix decreased by 70% after PAC treatment at the nano-length scale, while increased values were found at the micro-length scale (~0.24) compared to the control (0.18 ; p < .001). An increase in amide band intensities and a decrease of complex moduli was observed after storage in simulated body fluid for both Trimer-C and Trimer-EC modified dentin. The stereochemical configuration of the terminal monomeric units, C and EC, did not impact the chemo-mechanical stability of dentin matrix.

Discovery of Unusual Sesterterpenoids from Colquhounia coccinea var. mollis and Their Metabolic Implications.

Three unusual sesterterpenoids featuring unprecedented rearranged colquhounane (C25) and tetranorcolquhounane (C21) frameworks, colquhounoids E (1) and F (3) and norcolquhounoid F (2), were isolated from a Lamiaceae medicinal plant Colquhounia coccinea var. mollis. Their structures were elucidated by spectroscopic analysis and quantum chemical calculations. A biomimetic inspired regioselective cyclopropane cleavage was achieved under acidic conditions. The immunosuppressive activities of these new sesterterpenoids were also evaluated.

Unprecedented Benzoquinone Motifs Reveal Post-Oligomerizational Modification of Proanthocyanidins.

Proanthocyanidins (PACs) are complex flavan-3-ol polymers with stunning chemical complexity due to oxygenation patterns, oxidative phenolic ring linkages, and intricate stereochemistry of their heterocycles and inter-flavan linkages. Being promising candidates for dental restorative biomaterials, trace analysis of dentin bioactive cinnamon PACs now yielded novel trimeric (1 and 2) and tetrameric (3) PACs with unprecedented o- and p-benzoquinone motifs (benzoquinonoid PACs). Challenges in structural characterization, especially their absolute configuration, prompted the development of a new synthetic-analytical approach involving comprehensive spectroscopy, including NMR with quantum mechanics-driven 1H iterative functionalized spin analysis (HifSA) plus experimental and computational electronic circular dichroism (ECD). Vital stereochemical information was garnered from synthesizing 4-(2,5-benzoquinone)flavan-3-ols and a truncated analogue of trimer 2 as ECD models. Discovery of the first natural benzoquinonoid PACs provides new evidence to the experimentally elusive PAC biosynthesis as their formation requires two oxidative post-oligomerizational modifications (POMs) that are distinct and occur downstream from both quinone-methide-driven oligomerization and A-type linkage formation. While Nature is known to achieve structural diversity of many major compound classes by POMs, this is the first indication of PACs also following this common theme.

Labdane diterpenoids from the heartwood of Leucosceptrum canum that impact on root growth and seed germination of Arabidopsis thaliana.

Eleven undescribed diterpenoids possessing labdane, 3,18-cyclo-labdane, 19 (4 â†’ 3)-labdane and 12-nor-labdane skeletons, named leucolactones A-K, were isolated from the heartwood of a large woody Lamiaceae plant, Leucosceptrum canum. Their structures were determined by NMR, MS, and in the case of leucolactones A by single crystal X-ray diffraction analysis. Plausible biosynthetic pathway of leucolactones were proposed. Leucolactones showed significant inhibitory effects against seed germination and root elongation of Arabidopsis thaliana in the Petri dish bioassay. Among them, the diastereomeric leucolactones G and H were the most potent, with EC50 values for root elongation of 6.53 ± 1.35 and 9.75 ± 1.25 µM, respectively. The preliminary structure-activity relationship of leucolactones was discussed. The increase of auxin reporter activity in A. thaliana DR5::GUS roots by leucolactone H was observed, indicating that leucolactones altered auxin accumulation and distribution. These findings suggested that leucolactones might be involved in regulation of plant growth and development through altering auxin accumulation and distribution, presumably contributing to the heartwood formation in L. canum.

Seco B-Type Oligomers from Pinus massoniana Expand the Procyanidin Chemical Space and Exhibit Dental Bioactivity.

Investigation of a pine bark extract for bioactive proanthocyanidin oligomers resulted in the isolation of structurally related dimeric seco B-type procyanidin derivatives, 1-5. This includes scalemic mixtures of gambiriin A1 (1a) and A2 (2a) and their newly described optical antipodes, ent-gambiriin A1 (1b) and ent-gambiriin A2 (2b), respectively, as well as a racemic mixture of the newly described (ent-)gambiriin A5 (3a/3b). Furthermore, the study now fully characterizes the previously reported optically pure dimers gambiriin B1 (4) and gambirflavan D1 (5), and characterized the novel seco B-type procyanidin trimer, 6 (gambirifuran C1). Thermal conversion of catechin in aqueous solution provided further evidence for the structures of 1-6 and led to the purification of semisynthetic 1a and 2a as well as additional dimers 7-10. Elucidating the structures of the natural dimers, 1-5, from comprehensive NMR and ECD data and synthetic evidence provided crucial reference points for establishing the structure of the seco B-type procyanidin trimer, 6. Serving as assigned building blocks, data from the dimers supported the 3D structural assignment of 6 based on NMR substituent chemical shift differences (s.c.s., syn. ΔδC) and component-based empirical ECD calculations. Within the newly characterized series of PAC-related molecules, 5 exhibited high dentin biomodification potential. In addition, considering the nomenclature issues and plausible biosynthetic pathways of this group of compounds led to a consolidated nomenclature of all currently known seco B-type procyanidins. These findings, thereby, expand the chemical space of bioactive catechin oligomers, which have promise as agents for the natural enhancement of dental biomaterials. Finally, the current knowledge of the chemical space of seco B-type procyanidin derivatives was compiled to the level of absolute configuration.

B-type Proanthocyanidins with Dentin Biomodification Activity from Cocoa (Theobroma cacao).

To enable translational studies, a scalable preparative isolation scheme was developed for underivatized cocoa (Theobroma cacao) proanthocyanidins (PACs), affording six all-B-type oligomeric PACs, including a new tetramer 4. Their structures, including absolute configuration, were unambiguously established by comprehensive spectroscopic and chemical methods. Evaluation of the PACs' dentin biomodification properties employed dynamic mechanical and infrared spectroscopic analyses in dentin bioassay models. PAC treatment enhanced the biomechanical strength of dentin by 5- to 15-fold compared to untreated dentin. Among the PAC agents, the pentamer, cinnamtannin A3 (6), led to the highest complex modulus value of 131 MPa, whereas the "branched" tetramer, 4, showed the lowest, yet still significant bioactivity. This study of specifically singly linked medium-length oligomeric PACs indicates that the linkage site is paramount in determining the potency of these PACs as dentin biomodifiers.

Proanthocyanidin Tetramers and Pentamers from Cinnamomum verum Bark and Their Dentin Biomodification Bioactivities.

To enable the further exploration of structure-activity relationships (SARs) of proanthocyanidins (PACs) with dentin biomodification abilities, Cinnamomum verum was selected for scaled-up purification of mixed A-/B-type, medium-size PAC oligomers. Sequential purification by centrifugal partition chromatography (CPC), Sephadex LH-20, and semiprep HPLC chromatography yielded four underivatized tetrameric (5-8) and two pentameric (9-10) PACs. Their unambiguous structural characterization involved extensive spectral and chemical degradation approaches to show that epicatechin units are connected by plant-specific combinations of doubly linked A- and singly linked B-type interflavanyl bonds. The biomechanical properties (via dynamic mechanical analysis) and physicochemical structure (via infrared spectroscopy) were assessed to evaluate the biomodification potency of PAC-treated collagen in a preclinical dentin model. This study revealed that (4→8) versus (4→6) bonds in PAC interflavan linkages have limited influence on biomechanical outcomes of dentin. By exhibiting a 25-fold increase in the complex modulus of treated dentin compared to control, aesculitannin E (5) was found to be the most potent PAC known to date for enhancing the mechanical properties of dentin in this preclinical model.

Immunosuppresive Sesterterpenoids and Norsesterterpenoids from Colquhounia coccinea var. mollis.

A pair of new C-14 epimeric sesterterpenoids, colquhounoid D (1) and 14-epi-colquhounoid D (2), and five degradation products featuring new C20 and C21 frameworks, norcolquhounoids A-E (3-7), were isolated from Colquhounia coccinea var. mollis. Their structures were elucidated by comprehensive spectroscopic analysis and single-crystal X-ray diffraction. Degradation of the C25 skeleton to the C21 skeleton was also achieved using aqueous NaIO4 and RuCl3. Compounds 1 and 2 showed significant immunosuppressive activity on the cytokine IFN-γ secretion of mouse splenocytes induced by anti-CD3/CD4 monoclonal antibodies, with IC50 of 8.38 and 5.79 µM, respectively, and compounds 5 and 6 were moderately active.

Peltate glandular trichomes of Colquhounia vestita harbor diterpenoid acids that contribute to plant adaptation to UV radiation and cold stresses.

Plant glandular trichomes (GTs) are adaptive epidermal structures that synthesize and accumulate diverse specialized metabolites well-known as defense chemicals against biotic attacks, but their roles against abiotic challenges including UV radiation and cold climates remain largely unexplored. Colquhounia vestita Wall is a Chinese-Himalayan Lamiaceae plant with dense peltate and capitate GTs on its leaf and stem surfaces under a scanning electron microscope. Three diterpenoid acids, including a clerodane 5-epi-hardwickiic acid and two labdanes polyalthic acid and E-communic acid, were identified from the peltate GTs of C. vestita through laser microdissection coupled with UPLC-MS/MS. Under UV radiation and cold stresses, the major GT component polyalthic acid increased the biomass of Arabidopsis thaliana seedlings and decreased their malondialdehyde content. Furthermore, polyalthic acid promoted photosynthetic efficiency and the expression of genes encoding peroxidative enzymes under UV radiation, and stimulated Ca2+ elevation and the expression of calmodulin binding transcription activator gene CAMTA3 and two downstream cold-responsive genes CBF3 and RD29A under cold stress. Therefore, polyalthic acid in GTs is likely to endow the plant with enhanced tolerance to UV radiation and cold stresses, which extends the current understanding of the function of GT compounds in plant adaptation to abiotic environments.

Non-volatile natural products in plant glandular trichomes: chemistry, biological activities and biosynthesis.

Covering: 1960s to end of August 2018 Plant glandular trichomes (GTs) are adaptive structures that are well known as "phytochemical factories" due to their impressive capacity to biosynthesize and store large quantities of specialized natural products. The natural products in GTs are chemically diverse and mostly function as defense chemicals, therefore GTs are frequently regarded as "the first defense line" of plants against biotic and abiotic stresses. More importantly, many GT natural products are commercially desirable, thanks to their significant biological activities, thus attracting extensive interest in their biosynthesis. Consequently, it is well known that plant GTs are not only important reservoirs of biologically active natural products but are also a valuable bank of novel biosynthetic genes and enzymes. The non-volatile or oxygenated natural products in plant GTs, which need longer biosynthetic pathways and more energy from the plants, are of particular interest due to their more extensive biological activities and high commercial value. This review mainly focuses on these non-volatile natural products in plant GTs, including their chemistry, biological activities and biosynthesis. The methods employed for investigating natural products and their biosynthesis in plant GTs are also comprehensively discussed.

Chemical profile and defensive function of the latex of Euphorbia peplus.

Plant latex is an endogenous fluid secreted from highly specialized laticifer cells and has been suggested to act as a plant defense system. The chemical profile of the latex of Euphorbia peplus was investigated. A total of 13 terpenoids including two previously unknown diterpenoids, (2S*,3S*,4R*,5R*,6R*,8R*,l1R*,13S*,14S*,15R*, 16R*)-5,8,15-triacetoxy-3-benzoyloxy-11,16-dihydroxy-9-oxopepluane and (2R*,3R*, 4S*,5R*,7S*,8S*,9S*,l3S*,14S*,15R*)-2,5,8,9,14-pentaacetoxy-3-benzoyloxy-15-hydroxy-7-isobutyroyloxyjatropha-6(17),11E-diene), ten known diterpenoids, and a known acyclic triterpene alcohol peplusol, were identified, using HPLC and UPLC-MS/MS analyses and through comparison with the authentic compounds isolated from the whole plant. The diterpenoids exhibited significant antifeedant activity against a generalist plant-feeding insect, the cotton bollworm (Helicoverpa armigera), with EC50 values ranging from 0.36 to 4.60 µg/cm2. In particular, (2R*,3R*,4S*,5R*,7S*,8S*,9S*,l3S*,14S*,15R*)-2,5,9,14-tetraacetoxy-3-benzoyloxy-8,15-dihydroxy-7-isobutyroyloxyjatropha-6(17),11E-diene and (2R*,3R*, 4S*,5R*,7S*,8S*,9S*,l3S*,14S*,15R*)-2,5,14-triacetoxy-3-benzoyloxy-8,15-dihydroxy-7-isobutyroyloxy-9-nicotinoyloxyjatropha-6(17),11E-diene had EC50 values of 0.36 and 0.43 µg/cm2, respectively, which were approximately 7-fold more potent than commercial neem oil (EC50 = 2.62 µg/cm2). In addition, the major peplusol showed obvious antifungal activity against three strains of agricultural phytopathogenic fungi, Rhizoctonia solani, Colletotrichum litchi and Fusarium oxysporum f. sp. niveum. The results indicated that terpenoids in the latex of E. peplus are rich and highly diversified, and might function as constitutive defense metabolites against insect herbivores and pathogens for the plant.

Drimane Sesquiterpenoids and Isochromone Derivative from the Endophytic Fungus Pestalotiopsis sp. M-23.

Three new drimane sesquiterpenoids (1-3) together with the known 2α-hydroxyisodrimeninol (4), and a new isochromone derivative (5), were obtained from the solid cultures of fungal strain Pestalotiopsis sp. M-23, an endophytic fungus isolated from the leaves of Leucosceptrum canum (Labiatae). Their structures were determined by comprehensive 1D and 2D NMR, and MS analyses. The metabolites were evaluated for their antibacterial activities, and compound 3 showed weak inhibitory activity against Bacillus subtilis.

A Geranylfarnesyl Diphosphate Synthase Provides the Precursor for Sesterterpenoid (C25) Formation in the Glandular Trichomes of the Mint Species Leucosceptrum canum.

Plant sesterterpenoids, an important class of terpenoids, are widely distributed in various plants, including food crops. However, little is known about their biosynthesis. Here, we cloned and functionally characterized a plant geranylfarnesyl diphosphate synthase (Lc-GFDPS), the enzyme producing the C25 prenyl diphosphate precursor to all sesterterpenoids, from the glandular trichomes of the woody plant Leucosceptrum canum. GFDPS catalyzed the formation of GFDP after expression in Escherichia coli. Overexpressing GFDPS in Arabidopsis thaliana also gave an extract catalyzing GFDP formation. GFDPS was strongly expressed in glandular trichomes, and its transcript profile was completely in accordance with the sesterterpenoid accumulation pattern. GFDPS is localized to the plastids, and inhibitor studies indicated its use of isoprenyl diphosphate substrates supplied by the 2-C-methyl-D-erythritol 4-phosphate pathway. Application of a jasmonate defense hormone induced GFDPS transcript and sesterterpenoid accumulation, while reducing feeding and growth of the generalist insect Spodoptera exigua, suggesting that these C25 terpenoids play a defensive role. Phylogenetic analysis suggested that GFDPS probably evolved from plant geranylgeranyl diphosphate synthase under the influence of positive selection. The isolation of GFDPS provides a model for investigating sesterterpenoid formation in other species and a tool for manipulating the formation of this group in plants and other organisms.

Capitate glandular trichomes of Paragutzlaffia henryi harbor new phytotoxic labdane diterpenoids.

The morphology and chemical profile of the capitate glandular trichomes (CGTs) of Paragutzlaffia henryi (Acanthaceae) were investigated. Four new labdane diterpenoids named paraguhenryisins A-D (1-4), together with the known physacoztomatin (5), were localized to the CGTs using laser microdissection coupled with cryogenic (1)H NMR and HPLC analyses and were traced and isolated from the CGT extract of inflorescences. Their structures were determined by spectroscopic methods and single-crystal X-ray diffraction. Bioassays indicated significant inhibitory effect for these diterpenoids on Arabidopsis thaliana seed germination and seedling root elongation. The most potent inhibitor, paraguhenryisin C (3), was interestingly detected in both the rhizosphere soil and water rinsed inflorescences extract of P. henryi but not the roots, with average contents in the rhizosphere soil relevant to its phytotoxic EC50 values. These results suggested that phytotoxic labdane diterpenoids in the CGTs might be released into the environment as a defensive measure for P. henryi against other competitive plants.

Macrocyclic Diterpenoids from the Latex of Euphorbia helioscopia.

One new jatrophane diterpenoid, 7α,9ß,15ß-triacetoxy-3ß-benzoyloxy-14ß-hydroxyjatropha-5E,11E-diene (3), together with four known macrocyclic diterpenoids, euphoheliosnoid A (1), epieuphoscopin B (2), euphohelioscopin A (4) and euphoscopin C (5), were isolated from the stem latex of Euphorbia helioscopia. Their structures were established by spectroscopic analyses. In the anti-inflammatory assay, euphohelioscopin A (4) exhibited moderate inhibitory activity on the release of cytokine TNF-α (IC50 = 23.7 ± 1.7 µM), IL-6 (IC50 = 46.1 ± 1.1 µM) and chemokine MCP-1 (IC50 = 33.7 ± 3.8 µM) in lipopolysaccharide (LPS) induced RAW 264.7 macrophages without notable cytotoxicity (IC50 > 80 µM).

Unraveling the metabolic pathway in Leucosceptrum canum by isolation of new defensive leucosceptroid degradation products and biomimetic model synthesis.

Seven new leucosceptroid degradation products possessing a C20, C21, or C25 framework, norleucosceptroids D-H (1-5), leucosceptroids P (6), and Q (7), have been isolated from Leucosceptrum canum. Their structures were determined by comprehensive NMR, MS, and single-crystal X-ray diffraction analyses. Discovery of these key intermediates, together with the biomimetic oxidation of a model system, supports the hypothesis that two biosynthetic pathways are operative. Antifeedant activity was observed for compounds 1-3.

Antifeedant and Antiviral Diterpenoids from the Fresh Roots of Euphorbia jolkinii.

ABSTRACT: The perennial herbaceous plant Euphorbia jolkinii (Euphorbiaceae) is a noxious weed widely distributed in the grasslands of northwestern Yunnan and has greatly threatened the local biodiversity. Phytochemical investigation on the fresh roots of E. jolkinii afforded six new diterpenoids 1, 2, 4-6, and 8, together with fifteen known diterpenoids. Their structures were elucidated on the basis of 1D and 2D NMR and other spectroscopic methods. Casbane, lathyrane, abietane, and ent-kaurane diterpenoids were reported from this plant for the first time. Selected compounds were evaluated for their antifeedant and anti-RSV (respiratory syncytial virus) activities. Compound 2 and ingenol (3) exhibited moderate antifeedant activity against a generalist insect herbivore, Spodoptera exigua, with EC50 values of 17.88 and 17.71 µg/cm(2) respectively. Compound 19 showed significant anti-RSV activity, with 50 % inhibition (IC50) value of 10.0 µM and selective index of 8.0. Compounds 1 and 2 were less active against RSV virus, both with IC50 value of 25 µM, and with selective indices of 1.0 and 3.2 respectively. These findings provided new evidence for the biological functions and utilization of the diversified diterpenoid metabolites in the roots of this rich but harmful plant.

Biologically active dichapetalins from Dichapetalum gelonioides.

A phytochemical investigation of the toxic tropical plant Dichapetalum gelonioides led to the isolation and identification of 14 new dichapetalins (1-14) and the known dichapetalins A (15) and K (16). The structures of the new compounds were determined by analyses of their NMR, MS, electronic circular dichroism, and X-ray diffraction data. The esterification at C-25 by 4-hydroxyphenylpropanoic acid and the hydroxylation at C-2' are unique in this unusual class of natural products. In addition to the known cytotoxicity, an array of biological activities, including antifeedant, nematicidal, antifungal, and NO and AChE inhibitory activities, were observed for this class of compounds. These findings suggested that dichapetalin hybrid triterpenoids as a class have broad biologically active cellular functions including defense against insect herbivores and pathogens.