Kolavenic Acid Research Articles

KIFC1 regulates ZWINT to promote tumor progression and spheroid formation in colorectal cancer.

Colorectal cancer (CRC) is the second leading cause of cancer-related mortality worldwide. Kinesin Family Member C1 (KIFC1) has been proposed as a promising therapeutic target due to its pivotal role in centrosome clustering to mediate cancer cell progression. This study aimed to analyze the expression and biological function of KIFC1 in CRC. Immunohistochemically, 67 (52%) of 129 CRC cases were positive for KIFC1 and statistically associated with poorer overall survival. KIFC1 small interfering RNA (siRNA)-transfected cells demonstrated lower cell proliferation as compared to the negative control cells. A specific KIFC1 inhibitor, kolavenic acid analog (KAA) drastically inhibited CRC cell proliferation. Microarray analysis revealed that KAA-treated CRC cells presented reduced ZW10 interacting kinetochore protein (ZWINT) expression as compared to control cells. Immunohistochemical analysis demonstrated that 61 (47%) of 129 CRC cases were positive for ZWINT and ZWINT expression was significantly correlated with KIFC1 expression. ZWINT-positive cases exhibited significantly worse overall survival. KIFC1 siRNA-transfected cells showed reduced ZWINT expression while ZWINT siRNA-transfected cells decreased cell proliferation. Both KIFC1 and ZWINT knockdown cells attenuated spheroid formation ability. This study provides new insights into KIFC1 regulating ZWINT in CRC progression and its potential as a therapeutic target.

Antiamoebic activity of plant-based natural products and their conjugated silver nanoparticles against Acanthamoeba castellanii (ATCC 50492)

Acanthamoeba spp. are the causative agent of Acanthamoeba keratitis and granulomatous amoebic encephalitis (GAE). The current options to treat Acanthamoeba infections have limited success. Silver nanoparticles show antimicrobial effects and enhance the efficacy of their payload at the specific biological targets. Natural folk plants have been widely used for treating diseases as the phytochemicals from several plants have been shown to exhibit amoebicidal effects. Herein, we used natural products of plant or commercial sources including quercetin (QT), kolavenic acid (PGEA) isolated from plant extracts of Polyalthia longifolia var pendula and crude plant methanolic extract of Caesalpinia pulcherrima (CPFLM) as antiacanthamoebic agents. Furthermore, these plant-based materials were conjugated with silver nanoparticles (AgNPs) to determine the effects of the natural compounds and their nanoconjugates against a clinical isolate of A. castellanii from a keratitis patient (ATCC 50492) belonging to the T4 genotype. The compounds were conjugated with AgNPs and characterized by using ultraviolet visible spectrophotometry and atomic force microscopy. Quercetin coated silver nanoparticles (QT-AgNPs) showed characteristic surface plasmon resonance band at 443 nm and the average size distribution was found to be around 45 nm. The natural compounds alone and their nanoconjugates were tested for the viability of amoebae, encystation and excystation activity against A. castellanii. The natural compounds showed significant growth inhibition of A. castellanii while QT-AgNPs specifically exhibited enhanced antiamoebic effects as well as interrupted the encystation and excystation activity of the amoebae. Interestingly, these compounds and nanoconjugates did not exhibit in vitro cytotoxic effects against human cells. Plant-based compounds and extracts could be an interesting strategy in development of alternative therapeutics against Acanthamoeba infections.

Kolavenic acid analog restores growth in HSET-overproducing fission yeast cells and multipolar mitosis in MDA-MB-231 human cells

Although cancer cells often harbor supernumerary centrosomes, they form pseudo-bipolar spindles via centrosome clustering, instead of lethal multipolar spindles, and thus avoid cell death. Kinesin-14 HSET/KIFC1 is a crucial protein involved in centrosome clustering. Accordingly, a compound that targets HSET could potentially inhibit cancer cell proliferation in a targeted manner. Here, we report three natural compounds derived from Solidago altissima that restored the growth of fission yeast cells exhibiting lethal HSET overproduction (positive screening), namely solidagonic acid (SA) (1), kolavenic acid analog (KAA: a stereo isomer at C-9 and C-10 of 6β-tigloyloxykolavenic acid) (2), and kolavenic acid (KA) (3). All three compounds suppressed fission yeast cell death and enabled reversion of the mitotic spindles from a monopolar to bipolar morphology. Compound 2, which exerted the strongest activity against HSET-overproducing yeast cells, also inhibited centrosome clustering in MDA-MB-231 human breast adenocarcinoma cells, which contained large numbers of supernumerary centrosomes. These natural compounds may be useful as bioprobes in studies of HSET function. Moreover, compound 2 is a prime contender in the development of novel agents for cancer treatment.

Chemical defense against insects in Heterotheca subaxillaris and three Orobanchaceae species using exudates from trichomes.

One of the roles of plant trichomes is thought to be reducing feeding damage from herbivores. Among trichomes, glandular trichomes play a role in chemical defense systems in plants by means of stored biologically active phytochemicals. These phytochemicals act as pest repellents. They show antimicrobial and insecticidal activities, and they have also been isolated and identified from wild plants. The Asteraceae species Heterotheca subaxillaris has many glandular trichomes on the leaf surface, and these contain sesquiterpene carboxylates, which show insect antifeedant activity. Because these sesquiterpene carboxylates are major constituents of glandular trichomes, they may act as a chemical defense in H. subaxillaris. The Orobanchaceae species Parentucellia viscosa also has many glandular trichomes on the leaf surface and produces an insect antifeedant clerodane-type diterpene, kolavenic acid, in these trichomes. Additionally, two other Orobanchaceae species, Bellardia trixago and Parentucellia latifolia, also have many glandular trichomes, but the constituents of these glandular trichomes did not show biological activities against test insects. However, the seco-labdane diterpene alcohol trixagol and its hemi-malonate were major constituents in B. trixago, and these terpenes may act as physical defenses against herbivores by interfering with feeding due to their viscosity. The secondary metabolites from glandular trichomes of H. subaxillaris and P. viscosa showed insect antifeedant activity, and these secondary metabolites were presumed to act as chemical defenses for these plant species. On the other hand, non-biologically active secondary metabolites produced by two other Orobanchaceae, B. trixago and P. latifolia, were presumed to act as physical defenses due to their viscosity. Defense systems such as these may be applicable to new crop breeding to enhance protection against insect pests. © 2019 Society of Chemical Industry.

Quantification of diterpene acids in Copaiba oleoresin by UHPLC-ELSD and heteronuclear two-dimensional qNMR

In this study, we present the quantitation of eight diterpene acids in the oleoresin of Copaifera reticulata Ducke by UHPLC-ELSD and quantitative HSQC (heteronuclear single quantum correlation spectroscopy). UHPLC was performed using reversed phase material and external calibration and showed RSD values of ≤ 3% (repeatability) and ≤ 4% (precision), and mean recovery rates of 91.2–104.8%. LOQs were determined with 10 and 20 μg/mL, and LODs with 4 and 8 μg/mL, respectively. For the qHSQC method, calibration curves of eight different NMR cross-peaks (furylic, endo- and exocyclic methine signals, exocyclic methylene and methyl signals) were established and normalized with dimethyl terephthalate, which served as internal standard. This approach allowed the direct quantification of four major and one minor diterpene, whereas simple calculation procedures led to the contents of the remaining minor compounds. Comparison with the results of the UHPLC assay showed good agreement for seven of the eight diterpene acids. In terms of precision, the qHSQC method was advantageous for the quantification of the three main compounds, whereas UHPLC-ELSD was superior in the determination of the minor components. In contrast to previous reports, kolavenic acid was identified as a major diterpene acid in the oleoresin of Copaifera reticulata, with amounts of 4.0 ± 0.3%.

Antibiofilm potential of 16-oxo-cleroda-3, 13(14) E-diene-15 oic acid and its five new γ-amino γ-lactone derivatives against methicillin resistant Staphylococcus aureus and Streptococcus mutans

The clerodane diterpenoids 16-oxo-cleroda-3, 13(14) E-diene-15 oic acid (1) and kolavenic acid (2) isolated from Polyalthia longifolia var. pendula (Linn.) were previously reported for their antimicrobial activity. Thus present study was designed to investigate the biofilm inhibiting potential of these diterpenoids (1-2) and five new lactone derivatives (3-7) of 1 against methicillin resistant Staphylococcus aureus (MRSA), Streptococcus mutans, Klebsiella pneumoniae and Proteus mirabilis. Compounds 1 and 3 at 10-20 μg/mL were found to be bacteriostatic and significantly reduced the biofilm formation and metabolically active cells of MRSA and S. mutans up to 90%. Parental diterpenoid (1) at 10 and 16 μg/mL significantly eradicated the preformed biofilm of both pathogens. Both the compounds also delayed acid production at minimum inhibitory (MIC) and sub-inhibitory concentrations (sub MIC). Florescence and scanning electron microscopy further confirms the biofilm inhibiting potential of compounds 1 and 3 and displayed disrupted biofilms at MIC and sub MIC levels. The gene expression of MRSA and S. mutans responsible for biofilm formation was also altered. Moreover, the observed anti-virulence properties and delayed bacterial growth after 10 min of exposure to the test compounds 1 and 3 make them a promising class of antibiofilm agents.

Effect of Kolavenic Acid on the Structure of Human Serum Albumin

The effect of Kolavenic acid (KA), an active component isolated from the genus Polyalthia, on the structure of human serum albumin (HSA) was investigated by fluorescence polarization, synchronous fluorescence, three-dimensional (3D) fluorescence, and absorption spectroscopy in combination with molecular modeling techniques under physiological conditions. The synchronous and absorption fluorescence spectra confirmed that KA has an effect on the microenvironment around HSA in aqueous solution. The two-dimensional (2D) and 3D fluorescence spectra showed that KA could quench the intrinsic fluorescence of HSA and make its conformation change. Fluorescence polarization measurements provided useful information on the relaxation time and aggregation behavior of the complex formed between HSA and KA, and indicated that the presence of KA caused changes in the fluidity and microviscosity of HSA. The binding constants and thermodynamic parameters for KA-HSA systems were obtained under different temperatures (298, 308, and 318 K). Molecular docking showed that the KA moiety bound to the hydrophobic cavity of HSA, and there were three hydrogen-bonding interactions between KA and the Lys195 and Asp451 residues. Fluorescent displacement measurements confirmed that KA bound to HSA at site II. In addition, the binding mechanism of KA and HSA was revealed by the physicochemical parameters at the molecular level. The results showed that the interaction between KA and HSA was strong, indicating that KA may be stored and transferred by serum albumin.

Antileishmanial and Antimicrobial Clerodane Diterpenes from Polyalthia longifolia

Polyalthia longifolia is an evergreen tree, native to India, used in Indian System of Medicine as febrifuge and tonic. The stem-bark extracts of this plant and its isolated compounds were reported previously for various biological activities, including antibacterial, antifungal and cytotoxic activities [1 – 3]. In our investigation, three clerodane diterpenes were isolated by bioassay-guided fractionation and identified as (-) kolavenic acid (1), 16α-hydroxycleroda-3,13(14)Z-diene-15,16-olide (2) and 16-oxocleroda-3,13(14)E-diene-15-oic acid (3), using 1H- and 13C NMR, as well as 2D NMR experiments. Compound 1 displayed antibacterial activity against Staphylococcus aureus and methicillin-resistant S. aureus (MRSA) with IC50 values of 8.32 and 5.08 µg/mL, respectively, while compound 2 showed very weak activity against MRSA (IC50 18.95 µg/mL). On the other hand, compound 2 and its acetylation product (4) displayed antileishmanial activity with IC50 values of 4.50 and 3.60 µg/mL, respectively, against Leishmania donovani promastigote. Both compounds, 2 and 4, were also evaluated against L. donovani axenic amastigote and amastigote in THP1 macrophage cultures, where they displayed strong activities with IC50 values of < 1.60 and 2.34 µg/mL, and 2.34 and 3.72 µg/mL, respectively. The anticancer activity of these compounds was also evaluated.

Clerodane Diterpenes Isolated from Polyalthia longifolia Induce Apoptosis in Human Leukemia HL-60 Cells

Polyalthia is a versatile genus of shrubs and trees found in tropic and sub-tropic regions. In this study, three clerodane diterpenes, kolavenic acid (1), polyalthialdoic acid (2), and 16α-hydroxy-cleroda-3,13(14)Z-dien-15,16-olide (3) isolated from Polyalthia longifolia leaves were evaluated for their apoptotic potential against human leukemia HL-60 cells. Compounds 2 and 3 inhibited cell proliferation with IC₅₀ values of 21.8 and 13.7 μM, respectively. Morphological changes and DNA fragmentation analysis indicated that these diterpenes induce apoptotic cell death in the HL-60 cells. Our results revealed the importance of P. longifolia as a chemopreventive medicinal plant.

An Unusual Bisnor-clerodane Diterpenoid from Polyalthia Simiarum

The stem bark of Polyalthia simiarum has yielded a new bisnor-type clerodane diterpenoid, 2-oxo-14,15-bisnor-3,11E-kolavadien-13-one (1), and three previously known clerodane derivatives, kolavenic acid (2), 16beta-hydroxycleroda-3,13(14)Z-dien-15,16-olide (3), and 16-oxocleroda-3,13(14)E-dien-15-oic acid (4). The structures of these compounds were unambiguously determined by extensive NMR studies as well as by comparison with related compounds. Till now this is the second report of the occurrence of any unusual C-18 clerodane diterpenoid from nature. The crude light petroleum extract and the purified compound 3 demonstrated moderate free radical scavenging activity with IC50 values of 21.5 and 23.5 microg/mL, respectively.

Simultaneous analysis of tissue- and genotype-specific variation in Solidago altissima (Asteraceae) rhizome terpenoids, and the polyacetylene dehydromatricaria ester

Solidago altissima is a dominant perennial of old field succession in North America with the ability to form dense, almost monospecific stands; the plant is also considered an aggressive invasive throughout Europe and Japan. S. altissima’s success is facilitated by large belowground rhizome systems that allow clonal spread and possible allelopathic suppression of competitors. A diversity of polar and nonpolar terpenes and polyacetylenes have been described from S. altissima rhizomes; however, there is little information documenting the concentration or distribution of these allelochemicals and how they relate to plant defense hypotheses. We thus developed a GC–MS method to simultaneously quantify rhizome terpenes and polyacetylenes that spanned a range of polarities and confirmed the presence of 19 terpenoids in addition to the polyacetylene dehydromatricaria ester (DME). We, (1) determined relative concentrations and variability of mono-, sesqui-, neutral diterpenes, diterpene acids and DME within a central NY population; (2) compared accumulation of these compounds in differently developed rhizome tissue; and (3) tested the alleopathic effect of DME at naturally occurring concentrations on germination and seedling growth of Asclepias syriaca, a common competitor of S. altissima. Overall, diterpene acids dominated the phytochemical profile of most genotypes (averaging 0.75% of fresh mass) with kolavenic acid being the single most abundant component. Monoterpene, sesquiterpene and acidic diterpene concentrations were all significantly greater in the actively elongating (current-year) rhizomes compared to established rhizomes and followed optimal defense theory predictions. Conversely, DME levels were lower in the current-year rhizomes than in established rhizomes. DME inhibited seed germination of A. syriaca in a dose-dependent manner. The tissue-specific accumulation of the different compounds may represent a biosynthetic and/or allocational strategy that limits the exposure of young-tissue to high concentrations of DME possibly limiting the cytotoxic effects of DME.

Ageing behaviour and analytical characterization of the Jatobá resin collected from Hymenaea stigonocarpa Mart.

This paper reports the results of an analytical study to characterize the trunk resin collected from the Hymenaea stigonocarpa Mart. species from the region of Minas Gerais (Brazil), popularly known as Jatobá resin. Hymenaea resins are reported to have been used in artistic applications such as protective varnishes in polychromed sculptures and paintings. Therefore, the identification of the main chemical changes that take place in the resin when it is prepared as a thin film exposed to atmospheric effects have been considered herein. Changes due to the degradation effect of light have been studied on a series of specimens prepared as a thin films and subjected to accelerated UV light ageing. The results based on GC–MS, THM–Py–GC–MS analyses, and on-line trimethylsilylation Py–GC–MS using hexamethyldislylazane as a derivatization reagent, have been compared. The study shows that eperuic acid and its Δ7 and Δ8 isomers, together with copalic acid, are the major components of the studied resin. Other compounds such as kolavenic acid, iso-ozic acid and epi-pinifolic acid are also present in this resin as minor components. This composition differs from those of resins obtained from other species of the Hymenaea genus growing in this Amazonian region. UV light ageing of the Jatobá resin prepared as a thin film results in the appearance of a new isomer of eperuic acid. Some changes in the relative content of the major components present in the resin have also been observed after ageing.

(−)-Kolavenic acid

In the two, almost identical, mol­ecules in the asymmetric unit of the title compound [systematic name: (E)-3-methyl-5-(1,2,4a,5-tetra­methyl-1,2,3,4,4a,7,8,8a-octa­hydro­naphthalen-1-yl)pent-2-enoic acid], C20H32O2, the rings are trans fused. The cyclo­hexane ring has a chair conformation and the cyclo­hexene ring a distorted half-boat conformation. The two independent mol­ecules are connected into a dimer via O—H⋯O hydrogen bonds. The dimers are associated into supra­molecular chains along c via C—H⋯O contacts.

Evaluation of Insecticidal Activity of Diterpenes and Lignans from Aristolochia malmeana against Anticarsia gemmatalis

The insecticidal activity of hexane extracts from the roots and leaves of Aristolochia malmeana was evaluated against Anticarsia gemmatalis larvae by topical application. Extract from the roots was the most active and caused 50% mortality in larvae at 308.4 microg/microL. From this extract, a clerodane diterpene, (-)-kolavenic acid, and three lignans, (-)-kusunokinin, (-)-hinokinin, and (8 S,8' R,9 S)-cubebin, were isolated by chromatography and partition procedures and then evaluated for their insecticidal activities either individually or in pairs. (-)-Kusunokinin showed higher activity against A. gemmatalis (LD10=9.3, LD50=230.1 microg/microL) than the crude extract, and its activity was dose-dependent, whereas the other constituents did not exhibit any significant activity. Together with (-)-kusunokinin and (-)-hinokinin, (-)-copalic acid, (-)-2-oxokolavenic acid, (-)- ent-6-beta-hydroxy-copalic acid, (8 R,8' R,9 R)- and (8 R,8' R,9 S)-cubebins, (-)-fargesin, and (-)-phillygenin were isolated from the hexane extract of the leaves. The compounds were identified on the basis of spectroscopic analysis.

Hypotensive activity and toxicology of constituents from root bark ofPolyalthia longifolia var.pendula

A defatted extract of Polyalthia longifolia var. pendula root bark (PRB) in 50% methanol showed a significant ability to reduce blood pressure. It caused a 22% and 47% fall in mean arterial blood pressure (MABP) in rats at doses of 3 mg/kg and 30 mg/kg, respectively. Compounds purified from this extract include kolavenic acid (3), clerodane (1) and its isomer (2), liriodenine (4), lysicamine (5) and bisclerodane imide (6) and its isomer (7). Of these, only kolavenic acid produced a 22% fall in MABP, at a dose of 30 mg/kg. PRB showed a decrease in blood pressure of normotensive and egg yolk induced hypertensive rats. The LD50 of PRB was determined as 100 mg/kg in mice.

Antifungal clerodane diterpenes from Macaranga monandra (L) Muell. et Arg. (Euphorbiaceae).

Hexane and ethyl acetate phases of the methanol extract of Macaranga monandra showed fungal growth inhibition of Colletotrichum acutatum, C. fragariae and C. gloeosporioides, Fusarium oxysporum, Botrytis cinerea, Phomopsis obscurans, and P. viticola. Bioassay-guided fractionation led to the isolation of two active clerodane-type diterpenes that were elucidated by spectroscopic methods (1D-, 2D-NMR, and MS) as kolavenic acid and 2-oxo-kolavenic acid. A 96-well microbioassay revealed that kolavenic acid and 2-oxo-kolavenic acid produced moderate growth inhibition in Phomopsis viticola and Botrytis cinerea.

New antimicrobial alkaloids from the roots of Polyalthia longifolia var. pendula.

Bioassay-guided isolation studies on the root extract of Polyalthia longifolia var. pendula possessing significant antibacterial activity led to the isolation of three new alkaloids pendulamine A (1), pendulamine B (2) and penduline (3) along with stigmasterol 3-O-beta-D-glucoside, allantoin, the known diterpenoid kolavenic acid, and the azafluorene alkaloid isoursuline. The structures of these compounds have been elucidated with the help of spectroscopic studies including 2D NMR experiments. Compounds 1 and 2, which are the only protoberberine alkaloids having a monosubstituted A ring with a hydroxy group at C-3, were found to be the active antibacterial principles of the roots. Their MIC values ranged between 0.02-20 microg against the tested bacteria.

Effects of Acetylenes and Terpenoids from Solidago altissima L. on Seed Germination

Four kinds of acetylenes and three kinds of terpenoids were isolated from Solidago altissima L. and characterized by MS, 1H-NMR and 13C-NMR spectroscopy as methyl (Z) -decaene-4, 6, 8-triynoate (Z-dehydromatricaria esther, 1), methyl 10- [(Z) -2-methyl-2-butenoyloxy] - (2Z, 8Z) -2, 8-decadiene-4, 6-diyoate (2), (4Z) -2, 4-decadiene-6, 8-diyn-4-olide (Z-dehydromatricaria lactone, 3), (4E) -2, 4-decadiene-6, 8-diyn-4-olide (E-dehydromatricaria lactone, 4), 13E, 7α-acetoxyl kolavenic acid (solidagonic acid, 5), kolavenol (6), and D : C-friedours-7-en-3β-ol (ilexol, 7). Bioassay of these compounds on the germination of lettuce seeds showed that compounds 1, 2, 3, 4, and 5 exhibited growth inhibitory activity, whereas compound 7 did not exhibit the activity.

Antimicrobial diterpenes fromPolyalthia longifolia var.pendulla (Annonaceae)

The antimicrobial activities of the petroleum ether extract of the stem bark of Polyalthia longifolia var. pendulla as well as the diterpenes isolated from it were studied. Among the diterpenoids, 16-oxocleroda-3, 13E-dien-15-oic acid (1) exhibited the highest activities against most of the bacteria and the kanamycin resistant fungal strains, Asperigillus fumigatus, Saccharromyces caulbequence and S. cerevaceae, Candida albicans and Hensila californica compared with those of kolavenic acid (2) and 16β-hydroxycleroda-3,13-dien-15,16-olide (3). The minimum inhibitory concentrations for all of these compounds were determined and were found to be 8–64 μg/mL.

Polyacetylenes and diterpenes from Solidago canadensis

Investigation of the roots of Solidago canadensis resulted in the isolation of seven clerodane-type diterpenes related to kolavenic acid, one of wh