Derivatives of Amaryllidaceae Alkaloid Ambelline as Selective Inhibitors of Hepatic Stage of Plasmodium berghei Infection In Vitro.

The incidence rate of malaria and the ensuing mortality prompts the development of novel antimalarial drugs. In this work, the activity of twenty-eight Amaryllidaceae alkaloids (1-28) belonging to seven different structural types was assessed, as well as twenty semisynthetic derivatives of the ß-crinane alkaloid ambelline (28a-28t) and eleven derivatives of the α-crinane alkaloid haemanthamine (29a-29k) against the hepatic stage of Plasmodium infection. Six of these derivatives (28h, 28m, 28n and 28r-28t) were newly synthesized and structurally identified. The most active compounds, 11-O-(3,5-dimethoxybenzoyl)ambelline (28m) and 11-O-(3,4,5-trimethoxybenzoyl)ambelline (28n), displayed IC50 values in the nanomolar range of 48 and 47 nM, respectively. Strikingly, the derivatives of haemanthamine (29) with analogous substituents did not display any significant activity, even though their structures are quite similar. Interestingly, all active derivatives were strictly selective against the hepatic stage of infection, as they did not demonstrate any activity against the blood stage of Plasmodium infection. As the hepatic stage is a bottleneck of the plasmodial infection, liver-selective compounds can be considered crucial for further development of the malaria prophylactics.

Amaryllidaceae Alkaloids from Clivia miniata (Lindl.) Bosse (Amaryllidaceae): Isolation, Structural Elucidation, and Biological Activity.

Clivia miniata (Amaryllidaceae) is an herbaceous evergreen flowering plant that is endemic to South Africa and Swaziland and belongs to one of the top-10 traded medicinal plants in informal medicine markets in South Africa. The species has been reported as the most important component of a traditional healer's pallet of healing plants. Eighteen known Amaryllidaceae alkaloids (AAs) of various structural types, and one undescribed alkaloid of homolycorine-type, named clivimine B (3), were isolated from Clivia miniata. The chemical structures of the isolated alkaloids were elucidated by a combination of MS, HRMS, 1D and 2D NMR techniques and by comparison with literature data. Compounds isolated in a sufficient quantity, and not tested previously, were evaluated for their in vitro acetylcholinesterase (AChE; E.C. 3.1.1.7) and butyrylcholinesterase (BuChE; E.C. 3.1.1.8) inhibition activities.

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.

Monoterpene indole alkaloids from Vinca minor L. (Apocynaceae): Identification of new structural scaffold for treatment of Alzheimer's disease.

One undescribed indole alkaloid together with twenty-two known compounds have been isolated from aerial parts of Vinca minor L. (Apocynaceae). 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. The NMR data of several alkaloids have been revised, corrected, and missing data have been supplemented. Alkaloids isolated in sufficient quantity were screened for their in vitro acetylcholinesterase (AChE; E.C. 3.1.1.7) and butyrylcholinesterase (BuChE; E.C. 3.1.1.8) inhibitory activity. Selected compounds were also evaluated for prolyl oligopeptidase (POP; E.C. 3.4.21.26), and glycogen synthase 3ß-kinase (GSK-3ß; E.C. 2.7.11.26) inhibition potential. Significant hBuChE inhibition activity has been shown by (-)-2-ethyl-3[2-(3-ethylpiperidinyl)-ethyl]-1H-indole with an IC50 value of 0.65 ± 0.16 µM. This compound was further studied by enzyme kinetics, along with in silico techniques, to reveal the mode of inhibition. This compound is also predicted to cross the blood-brain barrier (BBB) through passive diffusion.

Semisynthetic Derivatives of Selected Amaryllidaceae Alkaloids as a New Class of Antimycobacterial Agents.

The search for novel antimycobacterial drugs is a matter of urgency, since tuberculosis is still one of the top ten causes of death from a single infectious agent, killing more than 1.4 million people worldwide each year. Nine Amaryllidaceae alkaloids (AAs) of various structural types have been screened for their antimycobacterial activity. Unfortunately, all were considered inactive, and thus a pilot series of aromatic esters of galanthamine, 3-O-methylpancracine, vittatine and maritidine were synthesized to increase biological activity. The semisynthetic derivatives of AAs were screened for their in vitro antimycobacterial activity against Mycobacterium tuberculosis H37Ra and two other mycobacterial strains (M. aurum, M. smegmatis) using a modified Microplate Alamar Blue Assay. The most active compounds were also studied for their in vitro hepatotoxicity on the hepatocellular carcinoma cell line HepG2. In general, the derivatization of the original AAs was associated with a significant increase in antimycobacterial activity. Several pilot derivatives were identified as compounds with micromolar MICs against M. tuberculosis H37Ra. Two derivatives of galanthamine, 1i and 1r, were selected for further structure optimalization to increase the selectivity index.

Alkaloids of Zephyranthes citrina (Amaryllidaceae) and their implication to Alzheimer's disease: Isolation, structural elucidation and biological activity.

Twenty known Amaryllidaceae alkaloids of various structural types, and one undescribed alkaloid of narcikachnine-type, named narcieliine (3), have been isolated from fresh bulbs of Zephyranthes citrina. The chemical structures of the isolated alkaloids were elucidated by a combination of MS, HRMS, 1D and 2D NMR, and CD spectroscopic techniques, and by comparison with literature data. The absolute configuration of narcieliine (3) has also been determined. Compounds isolated in a sufficient quantity were evaluated for their in vitro acetylcholinesterase (AChE; E.C. 3.1.1.7), butyrylcholinesterase (BuChE; E.C. 3.1.1.8), and prolyl oligopeptidase (POP; E.C. 3.4.21.26) inhibition activities. Significant human AChE/BuChE (hAChE/hBuChE) inhibitory activity was demonstrated by the newly described alkaloid narcieliine (3), with IC50 values of 18.7 ± 2.3 µM and 1.34 ± 0.31 µM, respectively. This compound is also predicted to cross the blood-brain barrier (BBB) through passive diffusion. The in vitro data were further supported by in silico studies of 3 in the active site of hAChE/hBuChE.

Amaryllidaceae Alkaloids of Belladine-Type from Narcissus pseudonarcissus cv. Carlton as New Selective Inhibitors of Butyrylcholinesterase.

Thirteen known (1-12 and 16) and three previously undescribed Amaryllidaceae alkaloids of belladine structural type, named carltonine A-C (13-15), were isolated from bulbs of Narcissus pseudonarcissus cv. Carlton (Amaryllidaceae) by standard chromatographic methods. Compounds isolated in sufficient amounts, and not tested previously, were evaluated for their in vitro acetylcholinesterase (AChE; E.C. 3.1.1.7), butyrylcholinesterase (BuChE; E.C. 3.1.1.8) and prolyl oligopeptidase (POP; E.C. 3.4.21.26) inhibition activities. Significant human BuChE (hBUChE) inhibitory activity was demonstrated by newly described alkaloids carltonine A (13) and carltonine B (14) with IC50 values of 913 ± 20 nM and 31 ± 1 nM, respectively. Both compounds displayed a selective inhibition pattern for hBuChE with an outstanding selectivity profile over AChE inhibition, higher than 100. The in vitro data were further supported by in silico studies of the active alkaloids 13 and 14 in the active site of hBuChE.

Functionalized aromatic esters of the Amaryllidaceae alkaloid haemanthamine and their in vitro and in silico biological activity connected to Alzheimer's disease.

A novel series of aromatic esters (1a-1m) related to the Amaryllidaceae alkaloid (AA) haemanthamine were designed, synthesized and tested in vitro with particular emphasis on the treatment of neurodegenerative diseases. Some of the synthesized compounds revealed promising acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) inhibitory profile. Significant human AChE (hAChE) inhibition was demonstrated by 11-O-(3-nitrobenzoyl)haemanthamine (1j) with IC50value of 4.0 ± 0.3 µM. The strongest human BuChE (hBuChE) inhibition generated 1-O-(2-methoxybenzoyl)haemanthamine (1g) with IC50 value 3.3 ± 0.4 µM. Moreover, 11-O-(2-chlorbenzoyl)haemanthamine (1m) was able to inhibit both enzymes in dose-dependent manner. The mode of hAChE and hBuChE inhibition was minutely inspected using enzyme kinetic analysis in tandem with in silico experiments, the latter elucidating crucial interaction in 1j-, 1m-hAChE and 1g-, 1m-hBuChE complexes. The blood-brain barrier (BBB) permeability was investigated applying the parallel artificial membrane permeation assay (PAMPA) to predict the CNS availability of the compounds.

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.

Amaryllidaceae Alkaloids as Potential Glycogen Synthase Kinase-3ß Inhibitors.

Glycogen synthase kinase-3ß (GSK-3ß) is a multifunctional serine/threonine protein kinase that was originally identified as an enzyme involved in the control of glycogen metabolism. It plays a key role in diverse physiological processes including metabolism, the cell cycle, and gene expression by regulating a wide variety of well-known substances like glycogen synthase, tau-protein, and ß-catenin. Recent studies have identified GSK-3ß as a potential therapeutic target in Alzheimer´s disease, bipolar disorder, stroke, more than 15 types of cancer, and diabetes. GSK-3ß is one of the most attractive targets for medicinal chemists in the discovery, design, and synthesis of new selective potent inhibitors. In the current study, twenty-eight Amaryllidaceae alkaloids of various structural types were studied for their potency to inhibit GSK-3ß. Promising results have been demonstrated by alkaloids of the homolycorine-{9-O-demethylhomolycorine (IC50 = 30.00 ± 0.71 µM), masonine (IC50 = 27.81 ± 0.01 µM)}, and lycorine-types {caranine (IC50 = 30.75 ± 0.04 µM)}.

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).

Anticancer potential of Amaryllidaceae alkaloids evaluated by screening with a panel of human cells, real-time cellular analysis and Ehrlich tumor-bearing mice.

In this study, twenty-two Amaryllidaceae alkaloids were screened for their anticancer potential. All isolates were evaluated for antiproliferative activities on a panel of 17 human cell types of different tissue origin using WST-1 assay. In addition, we determined the antiproliferative effect with a real-time cell analysis xCELLigence system. Thereafter, to evaluate the barely known in vivo anticancer potential of the most potent molecule haemanthamine, a preliminary study was performed using an Ehrlich tumor-bearing mice model. The results showed that haemanthamine, lycorine and haemanthidine exerted the highest antiproliferative activity. The mean growth percent (GP) value after a single-dose 10 µM treatment was for haemanthamine 21%, for lycorine 21% and for haemanthidine 27% that of untreated control cells (100%). Furthermore, haemanthamine, lycorine and haemanthidine exhibited significant cytotoxicities against all the tested cell lines with individual IC50 values in the micromolar range. Dynamic real-time measures of impedance by xCELLigence indicated that these three compounds suppress cell proliferation after 10 h of treatment at a concentration of 10 µM or higher. Regrettably, in a follow-up in vivo antitumor activity study, haemanthamine showed no statistically significant reduction in the tumor size with no prolongation of survival time of Ehrlich tumor-bearing mice. Taken together, these results provide a new clue and guidance for exploiting Amaryllidaceae alkaloids as anticancer agents.

AKR1C3 Inhibitory Potency of Naturally-occurring Amaryllidaceae Alkaloids of Different Structural Types.

Aldo-keto reductase 103 (AKRIC3) is an important human enzyme that participates in the reduction of steroids and prostaglandins, which leads to proliferative signaling. AKRIC3 is frequently upregulated in various cancers, and this enzyme has been suggested as a therapeutic target for the treatment of these pathological conditions. The fact that the isoquinoline alkaloid stylopine has been identified as a potent AKRIC3 inhibitor has prompted us to screen a library of diverse types of Amaryllidaceae alkaloids, which biogenetically are isoquinoline alkaloids, on a recombinant form of AKRIC3. From the tested compounds, only tazettine showed moderate AKRIC3 inhibitory potency with an IC5o value of 15.8 ? 1.2 pM. Tazettine is a common Amaryllidaceac alkaloid, which could be used as a model substance for the further development of either analogues or related compounds with better inhibition potency.

Cytotoxicity of Naturally Occurring Isoquinoline Alkaloids of Different Structural Types.

Forty-six isoquinoline alkaloids, of eleven structural types isolated in our laboratory, have been evaluated for their cytotoxicity against two cancer cell lines (Caco-2 and Hep-G2 cancer cells), as well as against normal human lung fibroblast cells. Only scoulerine, aromoline, berbamine and parfumidine showed significant cytotoxic effects, but only scoulerine was active against both Caco-2 and Hep-G2 cells (IC50 values 6.44 ± 0.87 and 4.57 ± 0.42, respectively). Unfortunately, except for parfumidine, the other active alkaloids were also cytotoxic to the normal human lung fibroblast cells.

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.

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.).

Antifungal and Antibacterial Activity of Extracts and Alkaloids of Selected Amaryllidaceae Species.

Alkaloidal extracts of six selected species of Amaryllidaceae were studied with respect to their antibacterial and anti-yeast activity and their alkaloidal fingerprint. Twenty-five alkaloids were determined by GC/MS, and sixteen of them identified from their mass spectra, retention times and retention indexes. In the antimicrobial assay, Gram-negative Escherichia coli and Gram-positive Staphylococcus aureus were used, along with isolates of the human pathogenic yeasts Candida albicans, C. glabrata, C. dubliniensis and Lodderomyces elongiosporus. The six extracts, together with 19 Amaryllidaceae alkaloids isolated in our laboratory, showed almost no inhibitory activity against the bacteria tested. However, promising anti-yeast properties were detected; the most potent activity was shown by lycorine, which inhibited C. dubliniensis with a MIC of 32 µg/mL, C. albicans and L. elongiosporus, both with MICs of 64 µg/mL, followed by caranine inhibiting C. dubliniensis with a MIC of 128 µg/mL. Among the alkaloidal extracts, Narcissus jonquilla cv. Baby Moon showed the most potent anti-yeast activity, with minimal and average MIC values of 128 and 192 µg/mL, respectively, followed by Leucojum aestivum, Narcissus poeticus var. recurvus and N. canaliculatus (average MICs 256, 267 and 299 µg/mL, respectively). The lowest MIC value among extracts was obtained for N. canaliculatus against L. elongiosporus (MIC 64 µg/mL).

In Vitro Inhibitory Effects of 8-O-Demethylmaritidine and Undulatine on Acetylcholinesterase and Their Predicted Penetration across the Blood-Brain Barrier.

Alzheimer's disease is the most common cause of dementia. Currently, acetylcholinesterase (AChE) inhibition is the most widely used therapeutic treatment. A large number of naturally occurring compounds have been found to inhibit AChE. In this report the mechanism of AChE inhibition of two Amaryllidaceae alkaloids, 8-O-demethylmaritidine (1) and undulatine (2), and their possible penetration across the blood-brain barrier have been studied. Both compounds act via a mixed inhibition mechanism. Based on the parallel artificial permeation assay (PAMPA) for the prediction of blood-brain barrier (BBB) penetration, only 2 should be able to cross the BBB by passive permeation.

Isoquinoline alkaloids as prolyl oligopeptidase inhibitors.

Prolyl oligopeptidase is a cytosolic serine peptidase that hydrolyses proline-containing peptides at the carboxy terminus of proline residues. It has been associated with schizophrenia, bipolar affective disorder, and related neuropsychiatric disorders and therefore may have important clinical implications. Thirty-one isoquinoline alkaloids of various structural types, previously isolated in our laboratory, were screened for their ability to inhibit prolyl oligopeptidase. Promising results have been showed by alkaloids californidine (IC50=55.6±3.5 µM), dihydrosanquinarine (IC50=99.1±7.6 µM), corypalmine (IC50=128.0±10.5 µM) and N-methyllaurotetanine (IC50=135.0±11.7 µM).

Chemical composition of bioactive alkaloid extracts from some Narcissus species and varieties and their biological activity.

Alkaloid extracts of eight Narcissus (Amaryllidaceae) species and varieties were studied with respect to their acetylcholinesterase (HuAChE) and butyrylcholinesterase (HuBuChE) inhibitory activity and alkaloid patterns. Thirty alkaloids were determined by GC/MS, and twenty-five of them identified from their mass spectra, retention times and retention indexes. Promising HuAChE inhibition activity was demonstrated by six Narcissus taxa and HuBuChE inhibition by N. jonquila cv. Double Campernelle and N. nanus cv. Elka with IC50 values of 24.1 +/- 1.9 microg/mL and 25.1 +/- 1.8 microg/mL, respectively. Two alkaloids were isolated in pure form using preparative TLC and identified as the galanthamine type alkaloid narwedine and the lycorine type alkaloid incartine. Both compounds were tested for their biological activity. They were considered inactive in HuAChE/HuBuChE assays, but showed promising prolyl oligopeptidase inhibition activities with IC50 values of 0.95 +/- 0.12 mM and 0.91 g 0.09 mM, respectively.