Benzyltetrahydroisoquinoline Alkaloids Research Articles

Network pharmacology-based mechanism analysis of dauricine on the alleviating Aβ-induced neurotoxicity in Caenorhabditis elegans

BackgroundDauricine (DAU), a benzyl tetrahydroisoquinoline alkaloid isolated from the root of Menispermum dauricum DC, exhibits promising anti-Alzheimer’s disease (AD) effects, but its underlying mechanisms remain inadequately investigated. This paper aims to identify potential targets and molecular mechanisms of DAU in AD treatment.MethodsNetwork pharmacology and molecular docking simulation method were used to screen and focus core targets. Various transgenic Caenorhabditis elegans models were chosen to validate the anti-AD efficacy and mechanism of DAU.ResultsThere are 66 potential DAU-AD target intersections identified from 100 DAU and 3036 AD-related targets. Subsequent protein-protein interaction (PPI) network analysis identified 16 core targets of DAU for anti-AD. PIK3CA, AKT1 and mTOR were predicted to be the central targets with the best connectivity through the analysis of “compound-target-biological process-pathway network”. Molecular docking revealed strong binding affinities between DAU and PIK3CA, AKT1, and mTOR. In vivo experiments demonstrated that DAU effectively reduced paralysis in AD nematodes caused by Aβ aggregation toxicity, downregulated expression of PIK3CA, AKT1, and mTOR homologues (age-1, akt-1, let-363), and upregulated expression of autophagy genes and the marker protein LGG-1. Simultaneously, DAU increased lysosomal content and enhanced degradation of the autophagy-related substrate protein P62. Thioflavin T（Th-T）staining experiment revealed that DAU decreased Aβ accumulation in AD nematodes. Further experiments also confirmed DAU’s protein scavenging activity in polyglutamine (polyQ) aggregation nematodes.ConclusionCollectively, the mechanism of DAU against AD may be related to the activation of the autophagy-lysosomal protein clearance pathway, which contributes to the decrease of Aβ aggregation and the restoration of protein homeostasis.

Synthesis and Evaluation of (Bis)benzyltetrahydroisoquinoline Alkaloids as Antiparasitic Agents.

Visceral leishmaniasis and Chagas disease are neglected tropical diseases (NTDs) that severely impact the developing world. With current therapies suffering from poor efficacy and safety profiles as well as emerging resistance, new drug leads are direly needed. In this work, 26 alkaloids (9 natural and 17 synthetic) belonging to the benzyltetrahydroisoquinoline (BI) family were evaluated against both the pro/trypomastigote and amastigote forms of the parasites Leishmania infantum and Trypanosoma cruzi, the causative agents of these diseases. These alkaloids were synthesized via an efficient and modular enantioselective approach based on Bischler-Napieralski cyclization/Noyori asymmetric transfer hydrogenation to build the tetrahydroisoquinoline core. The bis-benzyltetrahydroisoquinoline (BBI) alkaloids were prepared using an Ullmann coupling of two BI units to form the biaryl ether linkage, which enabled a comprehensive survey of the influence of BI stereochemistry on bioactivity. Preliminary studies into the mechanism of action against Leishmania mexicana demonstrate that these compounds interfere with the cell cycle, potentially through inhibition of kinetoplast division, which may offer opportunities to identify a new target/mechanism of action. Three of the synthesized alkaloids showed promising druglike potential, meeting the Drugs for Neglected Disease initiative (DNDi) criteria for a hit against Chagas disease.

The visualization of the spatial distribution of Cocculus orbiculatus based on air flow-assisted desorption electrospray ionization mass spectrometry imaging

Cocculus orbiculatus (C. orbiculatus), the root of plants belonging to the Menispermaceae family, has been extensively used to treat various diseases, including malaria and rheumatism. The main chemicals in these plants are alkaloids; however, the spatial distribution of these compounds within the plant roots remains undefined. This study aimed to visualize the spatial distribution of C. orbiculatus using air flow-assisted desorption electrospray ionization mass spectrometry imaging (AFADESI-MSI). In total, the spatial distribution of four aporphine alkaloids, five benzyltetrahydroisoquinoline alkaloids, six bisbenzylisoquinoline alkaloids, and one morphinane alkaloid in the cork layer, xylem, and ray of the root of C. orbiculatus was observed; the distribution characteristics of the different compounds in C. orbiculatus were significantly different. This study provides a visualized spatial distribution analysis method for the characterization of metabolites in the root tissue of C. orbiculatus and also provides valuable information for the specificity of the root of C. orbiculatus, which is beneficial for understanding its chemical separation, biosynthesis, and pharmacological activities.

Chemistry and biology of ent-morphinan alkaloids.

Morphinan alkaloids have attracted constant attention since the isolation of morphine by Sertürner in 1805. However, a group of 45 compounds possessing a complete ent-morphinan backbone can also be found in the literature. These compounds are related to the morphinandienone subgroup and display a substitution pattern which is different from the morphinans. In particular, these alkaloids could be substituted at position C-2 and C-8 either by a hydroxy function or a methoxy moiety. Four groups of ent-morphinan alkaloids can be proposed, the salutaridine, pallidine, cephasugine and erromangine series. Interestingly, the botanical distribution of the ent-morphinans is more widespread than for the morphinans and includes the Annonaceae, Berberidaceae, Euphorbiaceae, Fumariaceae, Hernandiaceae, Lauraceae, Menispermaceae, Monimiaceae, Papaveraceae, and Ranunculaceae families. To date, their exact mode of production remains elusive and their interplay with the biosynthetic pathway of other classes of benzyltetrahydroisoquinoline alkaloids, in particular aporphines, should be confirmed. Exploration of the biological and therapeutic potential of these compounds is limited to some areas, namely central nervous system (CNS), inflammation, cancer, malaria and viruses. Further studies should be conducted to identify the cellular/molecular targets in view of promoting these compounds as new scaffolds in medicinal chemistry.

Rhizobium rhizogenes infection in threatened Indian orchid Dendrobium ovatum mobilises ‘Moscatilin’ to enhance plant defensins

The present study illustrates the transformation ability of two wild-type bacterial strains of Rhizobium rhizogenes (MTCC 532 and MTCC 2364) on the embryogenic callus and callus-derived plantlets of a threatened Indian orchid, Dendrobium ovatum. Co-culture of the bacterium with the explants gave marginal hairy root phenotype that failed to multiply in the culture medium. Some primary and secondary metabolites were subdued in infected explants. Moscatilin, the stilbenoid active principle in D. ovatum, was found below the detection limit. The presence of two metabolites viz., Laudanosine, a benzyltetrahydroisoquinoline alkaloid and Lyciumin B, a cyclic peptide, were detected exclusively in the infected explants. The subjugated amino acids and phenolics in the infected plantlets were routed to produce phytoanticipins, and phenanthrenes, strengthening the defence mechanism in infected tissues. This research implies that the plant's defence mechanism activation could have prevented the extensive hairy root formation in the explants, even though nodulations and phenotype transitions were witnessed. Moscatilin has a structural resemblance with Resveratrol, a phytoalexin that combats bacterial and fungal pathogens. The study favours the possibility of Moscatlin being a precursor for phenanthrene compounds, thereby serving as a ‘phytoanticipin’ during the infection phase.

Guanábana (Annona muricata L): Origen, características, cosecha, Postcosecha, actividad antioxidante, actividad antinflamatoria y beneficios para la salud

The soursop (Annona muricataL) initially existed in the slopes of Peru, it has a physiology and nutritional composition that is very rich in proteins, flavonoids and antioxidants. It has unquantifiable properties of high use for the human being,which are extracted from the pulp, seed and leaves. In this set of reviews,we will mention the antioxidant, anti-inflammatory and anti-cancer properties, with the aim of publicizing the profitable consumption and use of this fruit. But without a doubt, the great anticancer power that soursop possesses is the most outstanding of it, a review of various which was carried out, which emphasize the quantification of anaceous, neurotoxic acetogenins and benzyltetrahydroisoquinoline alkaloids, in the same way it was raised in antioxidant activity, highlighting the presence of vitamins C and E, anthocyanins, carotenoids, flavonoids and other phenolic compounds, we can affirm that its intake is a contribution of great importance to health.

Rapid, on-site quantitative determination of higenamine in functional food using a time-resolved fluorescence microsphere test strip

Higenamine (HIG), a benzyltetrahydroisoquinoline alkaloid found naturally in plants, is classified as an S3 Prohibited Substance in the 2020 World Anti-Doping Agency (WADA) report. To avoid problems such as doping violations in competitive events, it is necessary to develop rapid and sensitive detection methods. In this study, a highly-sensitive anti-HIG monoclonal antibody (mAb) was prepared and a time-resolved fluorescent microsphere immunochromatographic test strip (TRFM-ICTS) was established for the rapid quantitative detection of HIG in functional foods. Under optimized conditions, the TRFM-ICTS was compared with colloidal gold immunochromatographic test strip (CG-ICTS), and the half-maximal inhibitory concentration (IC50) of TRFM-ICTS was 1.37 ng/mL. The spiked recoveries ranged from 86.4% to 105.3%, which was consistent with the results of liquid chromatography-tandem mass spectrometry (LC-MS/MS) in the detection of real functional food. Therefore, TRFM-ICTS can be a candidate method for doping monitoring in functional foods and a powerful tool for HIG quantification.

Seco-neferine A–F, three new pairs of benzyltetrahydroisoquinoline alkaloid epimers from Plumula Nelumbinis and their activity

Three new pairs of benzyltetrahydroisoquinoline (BIQ) alkaloid epimers, Seco-neferine A-F (1–6), were isolated from an EtOH extract of Plumula Nelumbinis. The structures of these compounds were identified by a combination of NMR, HR-ESI-MS, circular dichroism, UV spectroscopic analyses and specific rotations. The structure of compounds 1–6 possesses high similarity with neferine, because these three pairs of epimers have the same skeleton as neferine. Compounds 1,2 and 5,6 are open-loop compounds of position 1′ and 1 of neferine respectively. The H connects with position 2′ N of compounds 1,2 is replaced by methyl, forming the structure of compounds 3,4. Moreover, six compounds were tested for cytotoxicity against MDA-MB-231 breast cancer cell. Compound 6 displayed moderate inhibitory effects on breast cancer with IC50 of 38.96 μM, while compounds 2,3,4 show certain inhibitory effects.

Dauricine Attenuates Spatial Memory Impairment and Alzheimer-Like Pathologies by Enhancing Mitochondrial Function in a Mouse Model of Alzheimer's Disease.

Alzheimer's disease (AD) is characterized by extracellular amyloid plaques composed of β-amyloid (Aβ) and intracellular neurofibrillary tangles containing hyperphosphorylated tau protein. No effective therapy is available for this disease. In this study, we investigated the potential therapeutic effects of dauricine (DAU), a benzyl tetrahydroisoquinoline alkaloid, on AD, and found that DAU administration significantly improved cognitive impairments in 3xTg-AD mice by decreasing Aβ plaques and hyperphosphorylated tau and increasing the hippocampal ATP level. Proteomic and western blot analyses revealed that DAU treatment mainly modified the expression of proteins involved in mitochondrial energy metabolism, such as Aco2, Ndufs1, Cox5a, and SDHB, and that of synapse-related proteins such as Syn1 and Syn2. Pathway analysis revealed that DAU modulated the tricarboxylic acid cycle, synaptic vesicle cycle, glycolysis, and gluconeogenesis in 3xTg-AD mice. Our study suggests that DAU may be a potential drug for the treatment of AD.

HPLC-Based Activity Profiling for Antiprotozoal Compounds in Croton gratissimus and Cuscuta hyalina.

In a screening of Sudanese medicinal plants for antiprotozoal activity, the chloroform fractions obtained by liquid-liquid partitioning from ethanolic extracts of fruits of Croton gratissimus var. gratissimus and stems of Cuscuta hyalina Roth ex Schult. exhibited in vitro activity against axenically grown Leishmania donovani amastigotes. This antileishmanial activity was localized by HPLC-based activity profiling. Targeted preparative isolation afforded flavonoids 1–6, 3-methoxy-4-hydroxybenzoic acid (7), and benzyltetrahydroisoquinoline alkaloids laudanine (8) and laudanosine (9) from C. gratissimus, and pinoresinol (10), isorhamnetin (11), (-)-pseudosemiglabrin (12), and kaempferol (13) from C. hyalina. The antiprotozoal activity of 1–13 against L. donovani (axenic and intracellular amastigotes), Trypanosoma brucei rhodesiense (bloodstream forms), and Plasmodium falciparum (erythrocytic stages), and the cytotoxicity in L6 murine myoblast cells were determined in vitro. Quercetin-3,7-dimethylether (6) showed the highest activity against axenic L. donovani (IC50, 4.5 µM; selectivity index [SI], 12.3), P. falciparum (IC50, 7.3 µM; SI, 7.6), and T. b. rhodesiense (IC50, 2.4 µM; SI, 23.2). The congener ayanin (2) exhibited moderate antileishmanial (IC50, 8.2 µM; SI, 12.2), antiplasmodial (IC50, 7.8 µM; SI, 12.9), and antitrypanosomal activity (IC50, 11.2 µM; SI, 8.9). None of the compounds showed notable activity against the intramacrophage form of L. donovani.

ELISA for the Detection of the Prohibited Doping Agent Higenamine.

Higenamine is a natural benzyltetrahydroisoquinoline alkaloid produced by various plants. In the World Anti-Doping Agency report of 2020, higenamine is classified as a class S3 (selective and nonselective β2-agonist) prohibited substance. To minimize the problems resulting from the misuse of higenamine-containing products as well as from the abuse of doping agents in sport, numerous higenamine-detection methods have been investigated. In the present study, a monoclonal antibody against the (S)-enantiomer of higenamine was successfully produced and applied in the indirect competitive ELISA to detect the content of (S)-higenamine in plant samples and related products. By immunizing BALB/c mice with higenamine-BSA, the aforementioned monoclonal antibody was produced even when the hapten number, which was the higenamine molecules conjugated to the BSA molecule, was relatively low (approximately 4). The MAb was characterized and utilized in the established icELISA assay with a detectable range of 7.81 - 125 ng/mL. The assay limit of detection (LOD) was 4.41 ng/mL, indicating higher sensitivity than the conventional HPLC-UV methods. Various validation processes demonstrated that icELISA was precise, with the maximum CV (%) of the intra- and inter-assays of 11.58% and 10.18%, respectively. Moreover, the assay was accurate, with the recovery rates of spiked (S)-higenamine ranging from 82% to 113%, and sufficiently reliable for the detection of (S)-higenamine in various kinds of samples. Notably, the present study describes the first immunoassay for (S)-higenamine.

ASARONE-DERIVED PHENYLPROPANOIDS AND ISOQUINOLINE-DERIVED ALKALOIDS FROM THE BARK OF Duguetia pycnastera (Annonaceae) AND THEIR CYTOTOXICITIES

The phytochemical investigation of the hexane and methanol extracts from the bark of Duguetia pycnastera Sandwith (Annonaceae) afforded seven known compounds, two asarone-derived phenylpropanoids and five isoquinoline-derived alkaloids. The asarones, γ-asarone (1-allyl-2,4,5-trimethoxybenzene) and 2,4,5-trimethoxy-styrene were isolated of the hexane extract while the aporphine alkaloids, O-methylmoschatoline, lysicamine, nornuciferidine, and guatterine N-oxide, and the benzyltetrahydroisoquinoline alkaloid, (S)-reticuline were isolated of the alkaloid fraction of the methanol extract. This is the first report of these compounds in D. pycnastera. γ-Asarone is being reported for the first time in the Annonaceae. Nornuciferidine is described for the second time in the Annonaceae while guatterine N-oxide is the third register. The structures of the isolated compounds were established by extensive analyses using 1D and 2D NMR spectroscopy in combination with MS. The cytotoxic activity of the isolated compounds (except for nornuciferidine) was evaluated against cancer and non-cancerous cell lines, in which lysicamine was the most active compound, mainly against HL-60, HepG2, and K562 with IC50 values of 24.40, 28.86 and 38.75 µmol L-1, respectively.

Exploring the Leaves of Annona muricata L. as a Source of Potential Anti-inflammatory and Anticancer Agents.

The use of anti-inflammatory natural products to treat inflammatory disorders for cancer prevention and therapy is an appealing area of interest in the last decades. Annona muricata L. is one of the many plant extracts that have been explored owing to their anti-inflammatory and anticancer effects. Different parts of A. muricata especially the leaves have been used for various ethnomedicinal purposes by traditional healers to treat several diseases including cancer, inflammation, diabetes, liver diseases, and abscesses. Some of these experience-based claims on the use of the plant have been transformed into evidence-based information by scientific investigations. The leaves of the plant have been extensively investigated for its diverse pharmacological aspects and found eminent for anti-inflammatory and anticancer properties. However, most studies were not on the bioactive isolates which were responsible for the activities but were based on crude extracts of the plant. In this comprehensive review, all significant findings from previous investigations till date on the leaves of A. muricata, specifically on their anti-inflammatory and anticancer activities have been compiled. The toxicology of the plant which has been shown to be due to the presence of neurotoxic annaceous acetogenins and benzyltetrahydro-isoquinoline alkaloids has also been updated to provide recent information on its safety aspects. The present knowledge of the plant has been critically assessed, aimed at providing direction toward improving its prospect as a source of potential anti-inflammatory and anticancer agents. The analysis will provide a new path for ensuring research on this plant to discover new agents to treat inflammatory diseases and cancer. Further in vitro and in vivo studies should be carried out to explore the molecular mechanisms underlying their anti-inflammatory responses in relation to anticancer activity and more detail toxicity study to ensure they are safe for human consumption. Sufficient preclinical data and safety data generated will allow clinical trials to be pursued on this plant and its bioactive compounds.

The Isoquinoline Alkaloid Dauricine Targets Multiple Molecular Pathways to Ameliorate Alzheimer-Like Pathological Changes In Vitro.

Alzheimer's disease (AD), the most common neurodegenerative disease, has no effective treatment. Dauricine (DAU), a benzyl tetrahydroisoquinoline alkaloid isolated from the root of Menispermum dauricum DC, reportedly has neuroprotective effects in cerebral ischemia. Here, we investigated the effects of DAU on N2a cells stably transfected with Swedish mutant amyloid precursor protein (N2a/APP), an AD-like cell model. ELISA and Western blot analysis revealed that DAU inhibited APP processing and reduced Aβ accumulation. In addition, DAU ameliorated tau hyperphosphorylation via PP2A, p35/25, and CDK5 pathways in N2a/APP cells. The amelioration of tau hyperphosphorylation by DAU was also validated in HEK293/Tau cells, another cell line with tau hyperphosphorylation. Proteomic analysis revealed 85 differentially expressed proteins in the lysates between the wild-type N2a cells (N2a/WT) and the N2a/APP cells in the presence or absence of DAU; these were classified into 6 main categories according to their functions: endoplasmic reticulum (ER) stress-associated proteins, oxidative stress-associated proteins, cytoskeleton proteins, molecular chaperones, mitochondrial respiration and metabolism-related proteins, and signaling proteins. Taken together, we demonstrated that DAU treatment reduces AD-like pathology, thereby suggesting that DAU has potential therapeutic utility in AD.

Alkaloids with antioxidant activities from Aconitum handelianum

A new C20-diterpenoid alkaloid handelidine (1) and twenty-seven known alkaloids (2–28) were isolated from the roots of Aconitum handelianum. Their structures were established on the basis of extensive spectroscopic analyses. The study indicated that denudatine-type C20-diterpenoid alkaloids with vicinal-triol system and benzyltetrahydroisoquinoline alkaloids exhibited significant antioxidant activities measured by three antioxidant test systems. The aconitine-type C19-diterpenoid alkaloids could serve as potential secondary antioxidants for their strong binding effects to metal ions.

In vitro opioid receptor affinity and in vivo behavioral studies of Nelumbo nucifera flower

Ethnopharmacological relevanceNelumbo nucifera Geartn., known as sacred lotus, has been used traditionally in South East Asia as a traditional medicine for various CNS disorders including stress, fever, depression, insomnia, and cognitive conditions. Aim of the studyTo investigate the in vitro cannabinoid and opioid receptor binding affinities, and in vivo behavioral actions of Nelumbo flower extracts and to isolate the potential compounds to treat CNS associated disorders. Materials and methodsThe white and pink flowers of N. nucifera were extracted with 95% EtOH, followed by acid–base partitioning using CHCl3 to give acidic and basic partitions. These partitions were subjected to Centrifugal Preparative TLC (CPTLC) to yield benzyltetrahydroisoquinoline (BTIQ) alkaloids and long chain fatty acids, identified by physical and spectroscopic methods. In addition, EtOH extracts and partitions were analyzed for chemical markers by UHPLC/MS and GC/MS. In vitro neuropharmacological effects were evaluated by cannabinoid (CB1 and CB2) and opioid [delta (δ), kappa (ĸ), and mu (µ)] competitive radioligand binding and GTPγS functional assays. The in vivo behavioral effect was studied through the use of the mouse tetrad assay at 10, 30, 75 and 100mg/kg/ip doses that revealed the effect on locomotion, catalepsy, body temperature, and nociception of acidic and basic CHCl3 partitions, fractions, and compounds. ResultsThree aporphines, nuciferine (1), N-nor-nuciferine (2), asimilobine (3), and five BTIQs, armepavine (4), O-methylcoclaurine (5), N-methylcoclaurine (6), coclaurine (7), neferine (10), and a mixture of linoleic and palmitic acids (LA and PA), were identified and evaluated for cannabinoid and opioid receptor displacement activities. Compounds 5–7 showed binding affinities for the ĸ opioid receptor with equilibrium dissociation constant (Ki) values of 3.5±0.3, 0.9±0.1, 2.2±0.2μM, respectively. Compound 10 displayed affinities for δ-and µ- opioid receptors with Ki values of 0.7±0.1 and 1.8±0.2μM, respectively, and was determined to be a weak δ agonist by GTPγS functional assay. The mixture of LA and PA (1:1) showed an affinity for δ opioid receptor with a Ki value of 9.2±1.1μM. The acidic and basic CHCl3 partitions, compounds 1 and 7, and 5–7 mixture were subjected to the tetrad assay, of which the acidic partition displayed decreased locomotion and increased catalepsy, antinociception, and hypothermia in animal at doses of 75–100mg/kg/ip, and also showed clonic-tonic seizures upon touch at 100mg/kg. ConclusionBioassay-guided isolation revealed compounds 5–7, 10, and the mixture of LA and PA displayed various degrees of opioid receptor radioligand displacement affinities. The in vivo tetrad assay of acidic CHCl3 partition, enriched with aporphines 1 and 2, displayed actions on all four points of behavioral parameters. It can be concluded that the in vivo mild canabimimetic-type effect observed for the CHCl3 partition is likely mediated through other CNS mechanisms since the extracts, partitions, and isolated compounds had no affinity for the in vitro CB1 and CB2 receptors. This work, along with traditional use and the reported bioactivities of the BTIQ alkaloids, suggested further studies on N. nucifera are needed to understand the roles that the extracts and/or individual compounds might contribute to the behavioral effects.

Novel N-oxide derivatives of benzyltetrahydroisoquinoline alkaloid from the tubers of Stephania viridiflavens H.S. Lo et M. Yang

An investigation on the phytochemistry of the medicinal plant Stephania viridiflavens H.S. Lo et M. Yang led to isolate two new naturally occurring benzyltetrahydroisoquinoline alkaloids, (+)-1S, 2R-laudanidine-Nβ-oxide 2 and (+)-1S, 2S-laudanidine-Nα-oxide 3, along with four known benzyltetrahydroisoquinoline alkaloids: (+)-laudanidine 1, (+)-reticuline 4, (+)-1S, 2R-reticuline-Nβ-oxide 5 and (+)-1S, 2S-reticuline-Nα-oxide 6. The structure and the stereochemistry of these compounds were determined on the basis of spectroscopic methods and also confirmed by partial synthesis. To examine putative acetycholinesterase (AChE) inhibitory or cytotoxic activities, various bioassays were performed, the N-oxide derivatives (5 and 6) demonstrated more potent cytotoxicity than the corresponding free base.

The Interaction of Telomere DNA G-Quadruplex with Threebis-Benzyltetrahydroisoquinoline Alkaloids

Telomeres are important multifunctional nucleoprotein structures located at the ends of eukaryotic chromosomes. Telomerase regulates telomere elongation, and its activity is associated with tumorigenesis. Because the activity of telomerase can be inhibited by G-quadruplex (G4) formation (a four-stranded DNA with stacks of G-quartets formed by four guanines in a planar structure), the role of G4 in cancer therapy has attracted many research interests. We studied the effects of three natural alkaloids-tetrandrine, fangchinoline, and berbamine-on the stability and formation of telomere DNA G4 with circular dichroism melting spectroscopy (melting-CD), variable temperature ultraviolet (melting-UV), proton nuclear magnetic resonance spectroscopy ((1)H NMR), and molecular docking, and examined the relationships among the alkaloid structure and their activities. We further investigated their cytotoxicity with the 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium bromide assay (MTT) and flow cytometry (FCM). The results demonstrated that alkaloids increased G4 stability and induced its formation, which added structure diversity of G4-ligands. The results showed that -OH at R(1), -OCH(3) at R(2), and [Formula: see text] at R(3) had higher stability than other substituent groups for these alkaloids. We also found a transition of antiparallel to parallel G4 as the temperature increased. The result indicated the possible advantage of parallel G4 in adversity. In addition, the alkaloids demonstrated a moderate cytotoxicity and proved to be cell cycle blocker in the G(1) phase. These alkaloids have revealed promising potentials to be the agents for antitumor therapy.

Alkaloids from Annona muricata leaves

We studied the alkaloid composition of Annona muricata L. (Annonaceae) leaves collected in February 2009 in Conakry Botanical Garden (Republic of Guinea). Ordinary CHCl3 extraction afforded total alkaloids (0.125%) that were separated into phenolic and nonphenolic parts. Column chromatography over silica gel of the nonphenolic fraction isolated three bases; of the phenolic fraction, one. Spectral data, physical constants of the alkaloids and their salts, chemical transformation, and comparison with authentic samples identified the pure alkaloids. Base 1, C17H15NO2, mp 120–122°C, mass spectrum (m/z): 265 [M] +, 264 [M – 1]+ (100), 250, 236, 235; identified as anonaine [1, 2]. Base 2, isolated as the hydrochloride, C19H19NO3·HCl, mp 244–246°C, identified as isolaureline [3, 4]. Base 3, C18H17NO3, mp 124–126°C (acetone), UV spectrum ( max, EtOH): 219, 282 (log 4.49, 4.25), spectrum similar to that of isolaureline. The mass spectrum of 3 had peaks for ions (m/z) 295 [M]+, 294 [M – 1]+, 280 [M – 15]+, 266 [M – 29]+, and 147.5 [M]++ that were characteristic of noraporphine alkaloids. A comparison of spectral data of 3 and 2 in addition to the difference in the molecular weights by 14 mass units suggested that 3 was norisolaureline. In fact, Hess methylation of 3 produced a base that was identical to 2. According to these results, 3 was identified as xylopine [2, 5]. Base 4, C17H19NO3, phenolic crystalline base, mp 218–220°C (acetone), identified as coclaurine [6, 7]. Thus, we studied the alkaloid composition of Annona muricata leaves and isolated for the first time and identified the aporphine alkaloids anonaine, isolaureline, and xylopine and the benzyltetrahydroisoquinoline alkaloid coclaurine, which was isolated previously from this plant [7].

β2-Adrenoceptor-Mediated Tracheal Relaxation Induced by Higenamine fromNandina domesticaThunberg

The fruit of Nandina domestica Thunberg (ND, Berberidaceae) has been used to improve cough and breathing difficulties in Japan for many years, but very little is known about the constituent of ND responsible for this effect. We have recently reported that the crude extract from ND (NDE) inhibits histamine- and serotonin-induced contraction of isolated guinea pig trachea, and the inhibitory activity was not explained by nantenine, a well-known alkaloid isolated from ND. To explore other constituent(s) of NDE with tracheal smooth muscle relaxant activity, we fractionated NDE and assessed the pharmacological effects of the fractions using isolated guinea pig tracheal ring preparations. NDE was introduced into a polyaromatic absorbent resin column and stepwise eluted to yield five fractions, among which only the 40 % methanol fraction was active in relaxing tracheal smooth muscle precontracted with histamine. Further separation of the 40 % methanol fraction with high-performance liquid chromatography yielded multiple subfractions, one of which was remarkably active in relaxing histamine-precontracted trachea. Chemical analysis with a time-of-flight mass spectrometer and nuclear magnetic resonance spectrometer identified the constituent of the most active subfraction as higenamine, a benzyltetrahydroisoquinoline alkaloid. The potency and efficacy of the active constituent from NDE in relaxing trachea were almost equivalent to synthetic higenamine. In addition, the effect of the active constituent from NDE was competitively inhibited by the selective beta (2)-adrenoceptor antagonist ICI 118,551. These results indicate that the major constituent responsible for the effect of NDE is higenamine, which probably causes the tracheal relaxation through stimulation of beta (2) adrenoceptors.