Alkaloids of Dicranostigma franchetianum (Papaveraceae) and Berberine Derivatives as a New Class of Antimycobacterial Agents.

Tuberculosis (TB) is a widespread infectious disease caused by Mycobacterium tuberculosis. The increasing incidence of multidrug-resistant (MDR) and extensively drug-resistant (XDR) strains has created a need for new antiTB agents with new chemical scaffolds to combat the disease. Thus, the key question is: how to search for new antiTB and where to look for them? One of the possibilities is to search among natural products (NPs). In order to search for new antiTB drugs, the detailed phytochemical study of the whole Dicranostigma franchetianum plant was performed isolating wide spectrum of isoquinoline alkaloids (IAs). The chemical structures of the isolated alkaloids were determined by a combination of MS, HRMS, 1D, and 2D NMR techniques, and by comparison with literature data. Alkaloids were screened against Mycobacterium tuberculosis H37Ra and four other mycobacterial strains (M. aurum, M. avium, M. kansasii, and M. smegmatis). Alkaloids 3 and 5 showed moderate antimycobacterial activity against all tested strains (MICs 15.625-31.25 µg/mL). Furthermore, ten semisynthetic berberine (16a-16k) derivatives were developed and tested for antimycobacterial activity. In general, the derivatization of berberine was connected with a significant increase in antimycobacterial activity against all tested strains (MICs 0.39-7.81 µg/mL). Two derivatives (16e, 16k) were identified as compounds with micromolar MICs against M. tuberculosis H37Ra (MIC 2.96 and 2.78 µM). All compounds were also evaluated for their in vitro hepatotoxicity on a hepatocellular carcinoma cell line (HepG2), exerting lower cytotoxicity profile than their MIC values, thereby potentially reaching an effective concentration without revealing toxic side effects.

Steroid Glycosides Hyrcanoside and Deglucohyrcanoside: On Isolation, Structural Identification, and Anticancer Activity.

Cardiac glycosides (CGs) represent a group of sundry compounds of natural origin. Most CGs are potent inhibitors of Na+/K+-ATPase, and some are routinely utilized in the treatment of various cardiac conditions. Biological activities of other lesser known CGs have not been fully explored yet. Interestingly, the anticancer potential of some CGs was revealed and thereby, some of these compounds are now being evaluated for drug repositioning. However, high systemic toxicity and low cancer cell selectivity of the clinically used CGs have severely limited their utilization in cancer treatment so far. Therefore, in this study, we have focused on two poorly described CGs: hyrcanoside and deglucohyrcanoside. We elaborated on their isolation, structural identification, and cytotoxicity evaluation in a panel of cancerous and noncancerous cell lines, and on their potential to induce cell cycle arrest in the G2/M phase. The activity of hyrcanoside and deglucohyrcanoside was compared to three other CGs: ouabain, digitoxin, and cymarin. Furthermore, by in silico modeling, interaction of these CGs with Na+/K+-ATPase was also studied. Hopefully, these compounds could serve not only as a research tool for Na+/K+-ATPase inhibition, but also as novel cancer therapeutics.

Chemistry and Biological Activity of Alkaloids from the Genus Lycoris (Amaryllidaceae).

Lycoris Herbert, family Amaryllidaceae, is a small genus of about 20 species that are native to the warm temperate woodlands of eastern Asia, as in China, Korea, Japan, Taiwan, and the Himalayas. For many years, species of Lycoris have been subjected to extensive phytochemical and pharmacological investigations, resulting in either the isolation or identification of more than 110 Amaryllidaceae alkaloids belonging to different structural types. Amaryllidaceae alkaloids are frequently studied for their interesting biological properties, including antiviral, antibacterial, antitumor, antifungal, antimalarial, analgesic, cytotoxic, and cholinesterase inhibition activities. The present review aims to summarize comprehensively the research that has been reported on the phytochemistry and pharmacology of the genus Lycoris.

Chemical and Biological Aspects of Montanine-Type Alkaloids Isolated from Plants of the Amaryllidaceae Family.

Plants of the Amaryllidaceae family are promising therapeutic tools for human diseases and have been used as alternative medicines. The specific secondary metabolites of this plant family, called Amaryllidaceae alkaloids (AA), have attracted considerable attention due to their interesting pharmacological activities. One of them, galantamine, is already used in the therapy of Alzheimer's disease as a long acting, selective, reversible inhibitor of acetylcholinesterase. One group of AA is the montanine-type, such as montanine, pancracine and others, which share a 5,11-methanomorphanthridine core. So far, only 14 montanine-type alkaloids have been isolated. Compared with other structural-types of AA, montanine-type alkaloids are predominantly present in plants in low concentrations, but some of them display promising biological properties, especially in vitro cytotoxic activity against different cancerous cell lines. The present review aims to summarize comprehensively the research that has been published on the Amaryllidaceae alkaloids of montanine-type.

Bersavine: A Novel Bisbenzylisoquinoline Alkaloidwith Cytotoxic, Antiproliferative and Apoptosis-Inducing Effects on Human Leukemic Cells.

Bersavine is the new bisbenzylisoquinoline alkaloid isolated from the Berberis vulgaris L.(Berberidaceae) plant. The results of cytotoxicity screening 48 h post-treatment showed thatbersavine considerably inhibits the proliferation and viability of leukemic (Jurkat, MOLT-4), colon(HT-29), cervix (HeLa) and breast (MCF-7) cancer cells with IC50 values ranging from 8.1 to 11 µM.The viability and proliferation of leukemic Jurkat and MOLT-4 cells were decreased after bersavinetreatment in a time- and dose-dependent manner. Bersavine manifested concentration-dependentantiproliferative activity in human lung, breast, ovarian and hepatocellular carcinoma cell linesusing a xCELLigence assay. Significantly higher percentages of MOLT-4 cells exposed to bersavineat 20 µM for 24 h were arrested in the G1 phase of the cell cycle using the flow cytometry method.The higher percentage of apoptotic cells was measured after 24 h of bersavine treatment. Theupregulation of p53 phosphorylated on Ser392 was detected during the progression of MOLT-4 cellapoptosis. Mechanistically, bersavine-induced apoptosis is an effect of increased activity ofcaspases, while reduced proliferation seems dependent on increased Chk1 Ser345 phosphorylationand decreased Rb Ser807/811 phosphorylation in human leukemic cells.

Amaryllidaceae Alkaloids of Different Structural Types from Narcissus L. cv. Professor Einstein and Their Cytotoxic Activity.

In this detailed phytochemical study of Narcissus cv. Professor Einstein, we isolated 23 previously known Amaryllidaceae alkaloids (1-23) of several structural types and one previously undescribed alkaloid, 7-oxonorpluviine. The chemical structures were identified by various spectroscopic methods (GC-MS, LC-MS, 1D, and 2D NMR spectroscopy) and were compared with literature data. Alkaloids which had not previously been isolated and studied for cytotoxicity before and which were obtained in sufficient amounts were assayed for their cytotoxic activity on a panel of human cancer cell lines of different histotype. Above that, MRC-5 human fibroblasts were used as a control noncancerous cell line to determine the general toxicity of the tested compounds. The cytotoxicity of the tested alkaloids was evaluated using the WST-1 metabolic activity assay. The growth of all studied cancer cell lines was inhibited by pancracine (montanine-type alkaloid), with IC50 values which were in the range of 2.20 to 5.15 µM.

The Genus Nerine Herb. (Amaryllidaceae): Ethnobotany, Phytochemistry, and Biological Activity.

Nerine Herbert, family Amaryllidaceae, is a genus of about 30 species that are native to South Africa, Botswana, Lesotho, Namibia, and Swatini (formerly known as Swaziland). Species of Nerine are autumn-flowering, perennial, bulbous plants, which inhabit areas with summer rainfall and cool, dry winters. Most Nerine species have been cultivated for their elegant flowers, presenting a source of innumerable horticultural hybrids. For many years, species of Nerine have been subjected to extensive phytochemical and pharmacological investigations, which resulted in either the isolation or identification of more than fifty Amaryllidaceae alkaloids belonging to different structural types. Amaryllidaceae alkaloids are frequently studied for their interesting biological properties, including antiviral, antibacterial, antitumor, antifungal, antimalarial, analgesic, cytotoxic, and cholinesterase inhibition activities. The present review aims to summarize comprehensively the research that has been reported on the phytochemistry and pharmacology of the genus Nerine.

Alkaloids from Narcissus poeticus cv. Pink Parasol of various structural types and their biological activity.

Fifteen Amaryllidaceae alkaloids (1-15) of various structural types were isolated by standard chromatographic methods from fresh bulbs of Narcissus poeticus cv. Pink Parasol. The chemical structures were elucidated by MS, and 1D and 2D NMR spectroscopic analyses, and by comparison with literature data. Narcipavline (5) and narcikachnine (6) are reported here for the first time. In their structure are combined two basic structural types of Amaryllidaceae alkaloids (galanthamine- and galanthindole-structural types), which represent a new structural type of these compounds. Alkaloids isolated in sufficient amounts were evaluated for their human erythrocytic acetylcholinesterase, and human serum butyrylcholinesterase (HuBuChE) inhibition activity using Ellman's method. Z-Gly-Pro-p-nitroanilide was used as substrate in the prolyl oligopeptidase (POP) assay. Untested alkaloids were also screened for their cytotoxic activity against a small panel of human cancer cells, which spanned cell lines from different tissue types. In parallel, MRC-5 human fibroblasts were employed to determine overall toxicity against noncancerous cells. Some compounds were evaluated for their antiprotozoal activity. The newly isolated alkaloid narcipavline (5) showed interesting HuBuChE inhibition activity (IC50 = 24.4 ± 1.2 µM), and norlycoramine (11) demonstrated promising POP inhibition (IC50 = 0.21 ± 0.01 mM).

Comparative cytotoxicity of chelidonine and homochelidonine, the dimethoxy analogues isolated from Chelidonium majus L. (Papaveraceae), against human leukemic and lung carcinoma cells.

BACKGROUND: The search for new anticancer compounds is a crucial element of natural products research. PURPOSE: In this study the effects of naturally occurring homochelidonine in comparison to chelidonine on cell cycle progression and cell death in leukemic T-cells with different p53 status are described. METHODS: The mechanism of cytotoxic, antiproliferative, apoptosis-inducing effects and the effect on expressions of cell cycle regulatory proteins was investigated using XTT assay, Trypan blue exclusion assay, flow cytometry, Western blot analysis, xCELLigence, epi-fluorescence and 3D super resolution microscopy. A549 cells were used for xCELLigence, clonogenic assay and for monitoring microtubule stability. RESULTS: We found that homochelidonine and chelidonine displayed significant cytotoxicity in examined blood cancer cells with the exception of HEL 92.1.7 and U-937 exposed to homochelidonine. Unexpectedly, homochelidonine and chelidonine-induced cytotoxicity was more pronounced in Jurkat cells contrary to MOLT-4 cells. Homochelidonine showed an antiproliferative effect on A549 cells but it was less effective compared to chelidonine. Biphasic dose-depended G1 and G2/M cell cycle arrest along with the population of sub-G1 was found after treatment with homochelidonine in MOLT-4 cells. In variance thereto, an increase in G2/M cells was detected after treatment with homochelidonine in Jurkat cells. Treatment with chelidonine induced cell cycle arrest in the G2/M cell cycle in both MOLT-4 and Jurkat cells. MOLT-4 and Jurkat cells treated with homochelidonine and chelidonine showed features of apoptosis such as phosphatidylserine exposure, a loss of mitochondrial membrane potential and an increase in the caspases -3/7, -8 and -9. Western blots indicate that homochelidonine and chelidonine exposure activates Chk1 and Chk2. Studies conducted with fluorescence microscopy demonstrated that chelidonine and homochelidonine inhibit tubulin polymerization in A549 cells. CONCLUSION: Collectively, the data indicate that chelidonine and homochelidonine are potent inducers of cell death in cancer cell lines, highlighting their potential relevance in leukemic cells.

In vitro immunomodulatory activity, cytotoxicity and chemistry of some central European polypores.

Context Some mushrooms of the order Polyporales are known for their immunomodulatory actions. Objective The objective of this study is to evaluate the in vitro phagocytic and cytotoxic effects of extracts from polyporales native to Central Europe. Materials and methods The effects of ethanol extracts from 27 polypore species on opsonized zymosan-induced phagocytosis of isolated human neutrophils were tested by a chemiluminescence method. Colon epithelial cell lines, Caco-2 and HT-29, were used for cytotoxicity assays, and extracts were chemically characterized in terms of total phenolic and ß-glucan content. Results We observed phagocytosis or respiratory burst enhancing activity in 17 extracts, of which five species, namely Aurantiporus fissilis (Berk. & M.A. Curtis) H. Jahn ex Ryvarden, Trametes gibbosa (Pers.) Fr., Piptoporus betulinus (Bull.) P. Karst, Neolentinus lepideus (Fr.) Redhead & Ginns, Polyporus squamosus (Huds.) Fr., significantly increased phagocytosis in granulocytes by 205, 181, 158, 155 and 141%, respectively. The ß-glucan content of the three most potent extracts was 58, 42 and 74 mg/g, respectively, and the polyphenol content was 155.6, 133.5 and 155.2 µmol of gallic acid equivalent/g, respectively. Some extracts showed cytotoxic activity, with higher cytotoxicity in Caco-2 than in HT-29 cells. Pycnoporus cinnabarinus (Jacq.) P. Karst. extract was cytotoxic to both cell lines, with IC50 values of 81 and 31 µg/mL, respectively. Discussion and conclusion The most promising extracts were from N. lepideus and Polyporus squamosus, which are edible species and may be considered safe. Our findings support their use as culinary preparations or food supplements for various immunological gut disorders.

Isoquinoline Alkaloids from Fumaria officinalis L. and Their Biological Activities Related to Alzheimer's Disease.

Two new isoquinoline alkaloids, named fumaranine (2) and fumarostrejdine (10), along with 18 known alkaloids were isolated from aerial parts of Fumaria officinalis. The structures of the isolated compounds were elucidated on the basis of spectroscopic analyses and by comparison with literature data. The absolute configuration of the new compound 2 was determined by comparing its circular dichroism spectra with those of known analogs. Compounds isolated in sufficient amounts were evaluated for their acetylcholinesterase, butyrylcholinesterase, prolyl oligopeptidase (POP), and glycogen synthase kinase-3ß inhibitory activities. Parfumidine (8) and sinactine (15) exhibited potent POP inhibition activities (IC50 99±5 and 53±2 µM, resp.).

The effect of Amaryllidaceae alkaloids haemanthamine and haemanthidine on cell cycle progression and apoptosis in p53-negative human leukemic Jurkat cells.

Plants from the Amaryllidaceae family have been shown to be a promising source of biologically active natural compounds of which some selected are currently in pre-clinical development. Regardless of interesting pioneer works, little is known about Amaryllidaceae alkaloids that have shown promising anti-cancer activities. The crinane group of the Amaryllidaceae, including haemanthamine and haemanthidine, was amongst the first of these compounds to exhibit an interesting cytotoxic potential against cancer cell lines. However, the mechanism of cytotoxic and anti-proliferative activity is not yet entirely clear. The primary objectives of the current study were to investigate the effects of haemanthamine and haemanthidine on the induction of apoptosis and the cell cycle regulatory pathway in p53-null Jurkat cells. Results indicate that haemanthamine and haemanthidine treatment decreases cell viability and mitochondrial membrane potential, leads to a decline in the percentage of cells in the S phase of the cell cycle, induces apoptosis detected by Annexin V staining and increases caspase activity. Dose dependent apoptosis was cross verified by fluorescence and bright field microscopy through Annexin V/propidium iodine staining and morphological changes which characteristically attend programmed cell death. The apoptotic effect of haemanthamine and haemanthidine on leukemia cells is more pronounced than that of gamma radiation. Contrary to gamma radiation, Jurkat cells do not completely halt the cell cycle 24h upon haemanthamine and haemanthidine exposure. Both Amaryllidaceae alkaloids accumulate cells preferentially at G1 and G2 stages of the cell cycle with increased p16 expression and Chk1 Ser345 phosphorylation. Concerning the pro-apoptotic effect, haemanthidine was more active than haemanthamine in the Jurkat leukemia cell line.

Analysis of Amaryllidaceae alkaloids from Zephyranthes grandiflora by GC/MS and their cholinesterase activity

Amaryllidaceae are known as ornamental plants, furthermore some species of this family contain galanthamine, an acetylcholinesterase inhibitor approved for the treatment of Alzheimer's disease, and other alkaloids with interesting pharmacological activity. The chemical composition of alkaloids from Zephyranthes grandiflora Lindl. was analyzed by GC/MS. Seven known compounds, belonging to five structural types of Amaryllidaceae alkaloids, were identified. The alkaloid extract from the bulbs showed promising cholinesterase inhibitory activities against human blood acetylcholinesterase (HuAChE; IC50 39.2±3.0 µg/mL) and human plasma butyrylcholinesterase (HuBuChE; IC50 356±9.3 µg/mL).

Free-radical scavenging activity of some European Polyporales.

The ethanol extracts of 30 mushroom species from the order Polyporales were tested for their free radical scavenging activities by means of the DPPH free-radical scavenging and hydrogen peroxide-induced luminol chemiluminescence assays. In the DPPH radical scavenging test, 7 species gave EC50 values of less than 1 mg/mL. The highest activity was exhibited by Fomitopsis pinicola (0.09 mg/mL). However, in the hydrogen peroxide-induced luminol chemiluminescence assay, only 4 species had EC50 values of less than 1 mg/mL, with the highest activity being exhibited also by F. pinicola (0.26 mg/mL). When compared with known antioxidants, all the mushroom extracts exhibited relatively low activity in both assays. The free radical scavenging results appeared not to be correlated with the total phenol contents of the extracts.

Condensed and hydrolysable tannins as antioxidants influencing the health.

Natural polyphenols are a wide class of secondary plant metabolites and represent an abundant antioxidant component of human diet. An important, but often neglected group of natural polyphenols, are tannins. This review offers a general description of chemistry of both hydrolysable and condensed tannins (proanthocyanidins), the mechanisms of their antioxidation action, like free radical scavenging activity, chelation of transition metals, inhibition of prooxidative enzymes and lipid peroxidation. The mechanisms of action of antibacterial, antiviral, anticarcinogenic, cardiovascular system preventing, and antiinflammatory effects as well as the absorption, metabolic fate and positive in vivo effects of tannins are enclosed.

Evaluation of natural antioxidants of Leuzea carthamoides as a result of a screening study of 88 plant extracts from the European Asteraceae and Cichoriaceae.

In recently, there has been a great interest in natural antioxidants as bioactive components of food, nutraceuticals or potential drugs against several diseases. In our study, 88 extracts from various parts of plants from European Asteraceae and Cichoriaceae were assayed for radical scavenging activity by means of DPPH (1,1-diphenyl-2-picryl hydrazyl radical) test using the SIA (Sequential injection analysis) method developed for this purpose in our laboratory. DPPH radical scavenging activity of all tested plant extracts was evaluated according to the IC(50) parameter. 29 extracts exhibited IC(50) value lower than 0.1 mg/mL. The leaves of Leuzea carthamoides (IC(50) = 0.046 mg/mL) were chosen as the most promising sample for a subsequent phytochemical study, which resulted in isolation of seven natural compounds, namely, 4',5,7-trihydroxy-6-methoxyflavone (hispidulin) (1), 5, 7, 3', 4'- tetrahydroxyflavanone (eriodictyol) (2), 3',4',5,7-pentahydroxy-6-methoxyflavonol (patuletin) (3), eriodictyol-7-beta-glucopyranoside (4), 6-hydroxykaempferol-7-O-(6''-O-acetyl-beta-D-glucopyranoside) (5), 4-hydroxybenzoic acid (6) and 3,4-dihydroxybenzoic acid (protocatechuic acid) (7). Antioxidant activity of the isolated compounds was evaluated by DPPH test and ferric reducing antioxidant power (FRAP) test and compared with trolox and quercetin. Both tests evaluated the flavonoid (5) as the most active antioxidant. This result was confirmed by comparison with known data concerning the structure/activity relationships of flavonoids.

In vitro antiplatelet activity of flavonoids from Leuzea carthamoides.

Plants and their secondary metabolites, including flavonoids, exhibit a wide range of biological effects. Consequently, natural substances are receiving an increased attention in medicinal research. Owing to these facts, in vitro antiplatelet activity of ethanol summary extract and four flavonoids from Leuzea carthamoides was determined in human platelet-rich plasma. Arachidonic acid (AA), adenosine diphosphate (ADP), collagen (COL), and thrombin were used as agonists of platelet aggregation. The summary extract showed a significant inhibition of the aggregation induced by COL and ADP. Of the tested flavonoids, eriodictyol (1) and patuletin (2) influenced COL- and AA-induced aggregation. Their IC(50) values are presented. Flavonoid glycosides eriodictyol-7-beta-glucopyranoside (3) and 6-hydroxykaempferol-7-O-(6''-O-acetyl-beta-D[small cap]-glucopyranoside) (4) were found to be weak antiplatelet agents. These results confirmed the fact that glucosylation decreases the antiplatelet activity. Quantitative composition of tested flavonoids in L. carthamoides extract was also determined. Though two of the tested flavonoids inhibited platelet aggregation, further evaluation of L. carthamoides, in order to discover other antiplatelet active compounds and possible adverse health effects, is needed.

Effect of some acetylcholinesterase reactivators on human platelet aggregation in vitro.

Acetylcholinesterase (AChE) reactivators are employed for the prophylaxis and treatment of intoxications with organophosphorus AChE inhibitors, including nerve agents and pesticides. For the recovery of inhibited enzyme, derivatives from the group of pyridinium or bispyridinium aldoximes (called oximes) are used. Adverse effects of these substances are not well elucidated, because of their narrow and one-shot usage. Owing to this fact, the study evaluated the influence of some currently applied oximes on human platelet aggregation in vitro. The antiplatelet activity of pralidoxime, obidoxime, HI-6, methoxime and HLö 7 was assayed in human platelet rich plasma (2.5 x 10(8) platelets.ml(-1)) at a concentration of 1.35 mM. Arachidonic acid (AA), adenosine diphosphate (ADP), collagen (COL) and thrombin (TR) were used as agonists of platelet aggregation. All tested substances, except pralidoxime and methoxime, caused a significant inhibition of the aggregation process induced by AA, ADP and COL. Of the oximes assayed, none was found to influence TR triggered aggregation. Since reduced platelet aggregation can play an important role as an adverse effect in reactivator administration, further evaluation is needed for the estimation of the real impact of active oximes to the aggregation process in humans.