Characterization of two CYP80 enzymes provides insights into aporphine alkaloid skeleton formation in Aristolochia contorta.

Aporphine alkaloids are a large group of natural compounds with extensive pharmaceutical application prospects. The biosynthesis of aporphine alkaloids has been paid attentions in the past decades. Here, we determined the contents of four 1-benzylisoquinoline alkaloids and five aporphine alkaloids in root, stem, leaf, and flower of Aristolochia contorta Bunge, which belongs to magnoliids. Two CYP80 enzymes were identified and characterized from A. contorta. Both of them catalyze the unusual C-C phenol coupling reactions and directly form the aporphine alkaloid skeleton. AcCYP80G7 catalyzed the formation of hexacyclic aporphine corytuberine. AcCYP80Q8 catalyzed the formation of pentacyclic proaporphine glaziovine. Kingdom-wide phylogenetic analysis of the CYP80 family suggested that CYP80 first appeared in Nymphaeales. The functional divergence of hydroxylation and C-C (or C-O) phenol coupling preceded the divergence of magnoliids and eudicots. Probable crucial residues of AcCYP80Q8 were selected through sequence alignment and molecular docking. Site-directed mutagenesis revealed two crucial residues E284 and Y106 for the catalytic reaction. Identification and characterization of two aporphine skeleton-forming enzymes provide insights into the biosynthesis of aporphine alkaloids.

Selective Activity Against Amastigote Forms of Trypanosoma cruzi and Leishmania infantum of Diastereomeric Dicentrine N-oxides.

As part of our continuous research for the discovery of bioactive compounds against Trypanosoma cruzi and Leishmania infantum, the alkaloid (6aS)-dicentrine (1) was oxidized to afford (6aS,6S)- (2) and (6aS,6R)- (3) dicentrine-N-oxides. Evaluation of the cytotoxicity against NCTC cells indicated that 2 and 3 are non-toxic (CC50>200 µM) whereas 1 demonstrated CC50 of 52.0 µM. Concerning T. cruzi activity against amastigotes, derivatives 2 and 3 exhibited EC50 values of 9.9 µM (SI>20.2) and 27.5 µM (SI>7.3), respectively, but 1 is inactive (EC50>100 µM). Otherwise, when tested against L. infantum amastigotes, 1 and 3 exhibited EC50 values of 10.3 µM (SI=5.0) and 12.7 µM (SI>15.7), respectively, being 2 inactive (EC50>100 µM). Comparing the effects of positive controls benznidazol (EC50=6.5 µM and SI>30.7) and miltefosine (EC50=10.2 µM and SI=15.2), it was observed a selective antiparasitic activity to diastereomers 2 and 3 against T. cruzi and L. infantum. Considering stereochemical aspects, it was suggested that the configuration of the new stereocenter formed after oxidation of 1 played an important role in the bioactivity against amastigotes of both tested parasites.

New aporphine alkaloids with antitumor activities from the roots of Thalictrum omeiense.

Two new aporphine alkaloids, 6aR-2'-(3-oxobutenyl)-thaliadin (1) and N-methylthalisopynine (2), along with ten known analogs (3-12), were isolated from the roots of Thalictrum omeiense W. T. Wang et S. H. Wang. Their structures were determined by extensive spectroscopic and X-ray crystallographic analyses. Compounds 1-7 and 9-12 were tested for their antiproliferative effects in vitro against two human cancer cell lines (A549 and MCF-7). Among them, compounds 1, 3, and 7 exhibited moderate inhibitory activity against the tested cell lines with IC50 values ranging from 23.73 to 34.97 µM.

Total Synthesis of the Racemate of Laurolitsine.

The total synthesis of laurolitsine was achieved for the first time. This reaction was accomplished in 14 steps with a 2.3% yield (this was calculated using 3-hydroxy-4-methoxybenzaldehyde as the starting material) starting from two simple materials, 3-hydroxy-4-methoxybenzaldehyde and 2-(3-hydroxy-4-methoxyphenyl)acetic acid, and the longest linear sequence consisted of 11 steps. The key steps included an electrophilic addition reaction in which a nitro group was reduced to an amino group using lithium tetrahydroaluminum and a Pd-catalyzed direct biaryl coupling reaction. In this paper, many of the experimental steps were optimized, and an innovative postprocessing method in which 2-(3-(benzyloxy)-4-methoxyphenyl)ethanamine is salted with oxalic acid was proposed.

Identification of the cytochrome P450s responsible for the biosynthesis of two types of aporphine alkaloids and their de novo biosynthesis in yeast.

Aporphine alkaloids have diverse pharmacological activities; however, our understanding of their biosynthesis is relatively limited. Previous studies have classified aporphine alkaloids into two categories based on the configuration and number of substituents of the D-ring and have proposed preliminary biosynthetic pathways for each category. In this study, we identified two specific cytochrome P450 enzymes (CYP80G6 and CYP80Q5) with distinct activities toward (S)-configured and (R)-configured substrates from the herbaceous perennial vine Stephania tetrandra, shedding light on the biosynthetic mechanisms and stereochemical features of these two aporphine alkaloid categories. Additionally, we characterized two CYP719C enzymes (CYP719C3 and CYP719C4) that catalyzed the formation of the methylenedioxy bridge, an essential pharmacophoric group, on the A- and D-rings, respectively, of aporphine alkaloids. Leveraging the functional characterization of these crucial cytochrome P450 enzymes, we reconstructed the biosynthetic pathways for the two types of aporphine alkaloids in budding yeast (Saccharomyces cerevisiae) for the de novo production of compounds such as (R)-glaziovine, (S)-glaziovine, and magnoflorine. This study provides key insight into the biosynthesis of aporphine alkaloids and lays a foundation for producing these valuable compounds through synthetic biology.

Pathway elucidation and microbial synthesis of proaporphine and bis-benzylisoquinoline alkaloids from sacred lotus (Nelumbo nucifera).

Sacred lotus (Nelumbo nucifera) has been utilized as a food, medicine, and spiritual symbol for nearly 3000 years. The medicinal properties of lotus are largely attributed to its unique profile of benzylisoquinoline alkaloids (BIAs), which includes potential anti-cancer, anti-malarial and anti-arrhythmic compounds. BIA biosynthesis in sacred lotus differs markedly from that of opium poppy and other members of the Ranunculales, most notably in an abundance of BIAs possessing the (R)-stereochemical configuration and the absence of reticuline, a major branchpoint intermediate in most BIA producers. Owing to these unique metabolic features and the pharmacological potential of lotus, we set out to elucidate the BIA biosynthesis network in N. nucifera. Here we show that lotus CYP80G (NnCYP80G) and a superior ortholog from Peruvian nutmeg (Laurelia sempervirens; LsCYP80G) stereospecifically convert (R)-N-methylcoclaurine to the proaporphine alkaloid glaziovine, which is subsequently methylated to pronuciferine, the presumed precursor to nuciferine. While sacred lotus employs a dedicated (R)-route to aporphine alkaloids from (R)-norcoclaurine, we implemented an artificial stereochemical inversion approach to flip the stereochemistry of the core BIA pathway. Exploiting the unique substrate specificity of dehydroreticuline synthase from common poppy (Papaver rhoeas) and pairing it with dehydroreticuline reductase enabled de novo synthesis of (R)-N-methylcoclaurine from (S)-norcoclaurine and its subsequent conversion to pronuciferine. We leveraged our stereochemical inversion approach to also elucidate the role of NnCYP80A in sacred lotus metabolism, which we show catalyzes the stereospecific formation of the bis-BIA nelumboferine. Screening our collection of 66 plant O-methyltransferases enabled conversion of nelumboferine to liensinine, a potential anti-cancer bis-BIA from sacred lotus. Our work highlights the unique benzylisoquinoline metabolism of N. nucifera and enables the targeted overproduction of potential lotus pharmaceuticals using engineered microbial systems.

An Overview of Chemistry, Kinetics, Toxicity and Therapeutic Potential of Boldine in Neurological Disorders.

Boldine is an alkaloid obtained from the medicinal herb Peumus boldus (Mol.) (Chilean boldo tree; boldo) and belongs to the family Monimiaceae. It exhibits a wide range of pharmacological effects such as antioxidant, anticancer, hepatoprotective, neuroprotective, and anti-diabetic properties. There is a dearth of information regarding its pharmacokinetics and toxicity in addition to its potential pharmacological activity. Boldine belongs to the aporphine alkaloid class and possesses lipophilic properties which enable its efficient absorption and distribution throughout the body, including the central nervous system. It exhibits potent free radical scavenging activity, thereby reducing oxidative stress and preventing neuronal damage. Through a variety of neuroprotective mechanisms, including suppression of AChE and BuChE activity, blocking of connexin-43 hemichannels, pannexin 1 channel, reduction of NF-κß mediated interleukin release, and glutamate excitotoxicity which successfully reduces neuronal damage. These results point to its probable application in reducing neuroinflammation and oxidative stress in epilepsy, Alzheimer's disease (AD), and Parkinson's disease (PD). Moreover, its effects on serotonergic, dopaminergic, opioid, and cholinergic receptors were further investigated in order to determine its applicability for neurobehavioral dysfunctions. The article investigates the pharmacokinetics of boldine and reveals that it has a low oral bioavailability and a short half-life, requiring regular dosage to maintain therapeutic levels. The review studies boldine's potential therapeutic uses and mode of action while summarizing its neuroprotective benefits.  Given the favorable results for boldine as a potential neurotherapeutic drug in laboratory animals, more research is required. However, in order to optimise its therapeutic potential, it must be more bioavailable with fewer detrimental side effects.

A novel oxoisoaporphine-type alkaloid from the rhizome of Menispermum dauricum.

A phytochemical investigation of Menispermum dauricum led to the isolation of five oxoisoaporphine-type alkaloids (1-5) and five aporphine-type alkaloids (6-10), including a novel oxoisoaporphine-type alkaloid: menispeimin A (1). Their structures were elucidated by spectroscopic studies including MS, 1 D and 2 D NMR, and confirmed by comparing with literature data. Among them, alkaloids 4-10 were obtained for the first time from Menispermum genus. Natural products 2, 4 and 6 exhibited significant cytotoxic activity against A549, Bel-7402 and MCF-7 cell lines.

N-Methyl Costaricine and Costaricine, Two Potent Butyrylcholinesterase Inhibitors from Alseodaphne pendulifolia Gamb.

Studies have been conducted over the last decade to identify secondary metabolites from plants, in particular those from the class of alkaloids, for the development of new anti-Alzheimer's disease (AD) drugs. The genus Alseodaphne, comprising a wide range of alkaloids, is a promising source for the discovery of new cholinesterase inhibitors, the first-line treatment for AD. With regard to this, a phytochemical investigation of the dichloromethane extract of the bark of A. pendulifolia Gamb. was conducted. Repeated column chromatography and preparative thin-layer chromatography led to the isolation of a new bisbenzylisoquinoline alkaloid, N-methyl costaricine (1), together with costaricine (2), hernagine (3), N-methyl hernagine (4), corydine (5), and oxohernagine (6). Their structures were elucidated by the 1D- and 2D-NMR techniques and LCMS-IT-TOF analysis. Compounds 1 and 2 were more-potent BChE inhibitors than galantamine with IC50 values of 3.51 ± 0.80 µM and 2.90 ± 0.56 µM, respectively. The Lineweaver-Burk plots of compounds 1 and 2 indicated they were mixed-mode inhibitors. Compounds 1 and 2 have the potential to be employed as lead compounds for the development of new drugs or medicinal supplements to treat AD.

Protective effects of Stephania pierrei tuber-derived oxocrebanine against LPS-induced acute lung injury in mice.

Acute lung injury and acute respiratory distress syndrome (ALI/ARDS) have high mortality rates. Though corticosteroids are commonly used for the treatment of these conditions, their efficacy has not been conclusively demonstrated and their use can induce various adverse reactions. Hence, the application of corticosteroids as therapeutic modalities for ALI/ARDS is limited. Meanwhile, the aporphine alkaloid oxocrebanine isolated from Stephania pierrei tubers has demonstrated anti-inflammatory efficacy in murine/human macrophage cell lines stimulated by lipopolysaccharide (LPS). Accordingly, the primary objectives of the present study are to investigate the anti-inflammatory effects of oxocrebanine on LPS-induced murine alveolar epithelial (MLE-12) cells and its efficacy against LPS-induced murine ALI. Results show that oxocrebanine downregulates the abundance of interleukin (IL)-1beta, IL-6, and inducible nitric oxide synthase, as well as the phosphorylation of nuclear factor-kappaB (NF-κB), stress-activated protein kinase (SAPK)/c-Jun N-terminal kinase (JNK), p38, protein kinase B (Akt), and glycogen synthase kinase-3beta signalling proteins in LPS-induced MLE-12 cells. Moreover, in a murine ALI model, oxocrebanine lowers lung injury scores and lung wet/dry weight ratios while reducing inflammatory cell infiltration. It also suppresses LPS-induced tumour necrosis factor-alpha and IL-6 in the bronchoalveolar lavage fluid and plasma. Moreover, oxocrebanine downregulates NF-κB, SAPK/JNK, p38, and Akt phosphorylation in the lung tissues of LPS-treated mice. Taken together, the foregoing results show that oxocrebanine provides significant protection against LPS-induced ALI in mice primarily by suppressing various inflammatory signalling pathways in alveolar epithelial cells and lung tissues. Hence, oxocrebanine might prove effective as an anti-inflammatory agent for the treatment of lung inflammation.

Isoquinoline alkaloids isolated from Glaucium corniculatum var. corniculatum and Glaucium grandiflorum subsp. refractum var. torquatum with bioactivity studies.

CONTEXT: The genus Glaucium Mill., one of the important Papaveraceae family plants, is rich in isoquinoline alkaloids and distributed worldwide. OBJECTIVE: Isolation and identification of bioactive alkaloids from Glaucium grandiflorum Boiss. & Huet. subsp. refractum (Nabelek) Mory var. torquatum (Cullen) Mory and G. corniculatum (L.) Rudolph var. corniculatum (Aslan 2012), and investigation of their antioxidant and anticholinesterase activities. MATERIALS AND METHODS: The aerial parts of each plant were dried, powdered, and percolated with methanol, then each extract was fractionated between 50% aqueous acetic acid and petroleum. Their aqueous acidic layer was adjusted to pH 7-8 with NH4OH and extracted with chloroform, the extract was subjected to CC separation and isolation. Structures of the isolated alkaloids were elucidated by 1D and 2D-NMR and mass spectral analyses. The alkaloid extracts and their pure alkaloids were tested for anti-cholinesterase (AChE and BuChE) and antioxidant (ABTS, CUPRAC, ß-carotene linoleic acid tests) activities in vitro. RESULTS: Methanol extracts of Glaucium grandiflorum subsp. refractum var. torquatum and G. corniculatum var. corniculatum afforded a novel compound glauciumoline and seven known isoquinoline alkaloids three of which have an aporphine-type and the other five have a protopine-type skeleton. Among them, trans-protopinium (7) and cis-protopinium (8) were isolated from a Glaucium species for the first time. Tertiary amine extracts (TAEs) of both plants showed very strong acetylcholinesterase inhibitory activity. The TAE of the plants also showed strong antioxidant activity while the isolated alkaloids showed no meaningful activity in the anticholinesterase and antioxidant tests. DISCUSSION AND CONCLUSIONS: Glaucium species are considered promising therapeutic agents in the treatment of Alzheimer's disease.

Structure-activity relationship study of 4,5-didehydroguadiscine, an aporphine alkaloid showing potent melanogenesis-inhibitory activity in B16 melanoma cells.

Although 4,5-didehydroguadiscine (12a), an alkaloid with potent melanogenesis-inhibitory activity isolated from Hornschuchia obliqua (Annonaceae), consists of an aporphine nucleus with an aromatized B-ring, to date, it has not been utilized as a template for structure-activity relationship (SAR) studies of pharmacological activities because of its exceptional structure. Accordingly, herein, five analogs (12b-12f) of 12a and five benzylisoquinoline analogs (13b-13f) lacking the C11a-C11b bond of 12b-12f were prepared. The inhibitory effects of 12b-12f and 13b-13f on melanogenesis in theophylline-stimulated B16 melanoma 4A5 cells were examined and compared with those of 12a. Melanogenesis-inhibitory activities of 12b-12f were the same as that of 12a, whereas the melanogenesis-inhibitory activities of 13b-13f were significantly inferior to those of 12a and 12b-12f. These results suggest that the C11a-C11b bond plays an essential role in the melanogenesis-inhibitory activities of 12a-12e.

Cholinesterase inhibitors assessment of aporphine alkaloids from Annona crassiflora and molecular docking studies.

Annona crassiflora Mart. is an endemic plant from Brazilian Cerrado (savanna) biome, commonly employed in traditional medicine to treat wounds, diarrhea, and scalp infections. The pulp of the fruits is edible and has a characteristic taste, being employed to prepare sweets like jam, cakes, and ice cream by the people who live in the region of the Cerrado, although the peels are discarded. In this way, the A. crassiflora fruit peels ethanol extract was prepared and subjected to liquid-liquid extraction, which yielded the alkaloidal fraction (CH2Cl2). Subjected to high-performance liquid chromatography separations, this fraction allowed the purification of the aporphine alkaloids stephalagine (1), liriodenine (2), and atherospermidine (3), that were structurally characterized by high-resolution mass spectrometry with electrospray ionization, and nuclear magnetic resonance spectroscopy analyses. Aporphine alkaloids are recognized for their acetylcholinesterase (AChE) inhibitory activity, an important target in Alzheimer's disease therapy. Thus, the ethanol extract, alkaloidal fraction, and compounds1,2,and3were evaluated for acetyl- and butyrylcholinesterase (BChE) inhibitory activities. Compound1(IC50 104.2 µmol L-1) exhibited better BChE inhibitory activity than the standard compound galanthamine (IC50 162.7 µmol L-1), while2had a comparable result(and IC50 167.3 µmol L-1). Furthermore, molecular docking was performed to predict the compound's binding mode to the human AChE at a molecular level. Semiempirical calculation results show that the enthalpy interaction energy (ΔHint) between AChE and BChE active sites and all ligands were favorable for both enzymes, with the ligands interacting even more strongly with AChE, corroborating with IC50 results.

Structural optimizations and bioevaluation of N-H aporphine analogues as Gq-biased and selective serotonin 5-HT2C receptor agonists.

The concept of subtype selectivity and functional bias has recently reshaped current GPCR drug discovery for G protein-coupled receptors. A series of new N-H aporphines with A-ring modifications have been synthesized, and their efficacy on 5-HT2 receptor activation was evaluated. SAR analysis led to the discovery of several more potent and selective 5-HT2C receptor agonists (e.g., 11b and 11f) with low nanomolar activity. Molecular docking analysis of this series of aporphines was in accordance with our SAR results. The functional selectivity of specific compounds was tested via both Gq-mediated calcium flux and ß-arrestin recruitment assays, which revealed that these compounds exhibited no ß-arrestin recruitment activity. Further ADMET study combined with behavioral assessment using a methamphetamine-induced hyperactivity model identified compound 11b and 11f possessing promising drug-like profiles and having antipsychotic potential. These agonists with an exclusive bias toward Gq signaling may serve as valuable pharmacological probes to facilitate the elucidation of therapeutically relevant 5-HT2C signaling pathways and the development of alternative antipsychotic medications.

First Total Syntheses of Natural Phenanthrene Alkaloids, Uvariopsamine, Noruvariopsamine, 8-Hydroxystephenanthrine, 8-Methoxyuvariopsine, Thalihazine, and Secophoebine, and Their Potential as Antimalarial Agents.

The first total syntheses of natural phenanthrene alkaloids, namely, uvariopsamine (1), noruvariopsamine (2), 8-hydroxystephenanthrine (3), 8-methoxyuvariopsine (4), thalihazine (5), and secophoebine (6), have been realized. In addition, their in vitro antimalarial activity against the multidrug-resistant K1 strain of Plasmodium falciparum and in vitro cytotoxic activity against the human nasopharynx carcinoma (KB), small cell lung cancer (NCI-H187), and breast cancer (MCF7) human cancer cell lines were investigated. All the phenanthrene alkaloids showed significant antiplasmodial activity (IC50 1.07-7.41 µM), and most compounds displayed low to no toxicity against the three cancer cell lines tested. Particularly, 3 exhibited the best antimalarial activity with an IC50 value of 1.07 µM, no toxicity to NCI-H187 (IC50 > 50 µM), and low toxicity against KB (IC50 24.53 µM) and MCF7 (IC50 42.67 µM) cell lines.

Pharmacokinetics of glaucine after intravenous and oral administrations and detection of systemic aporphine alkaloids after ingestion of tulip poplar shavings in horses.

Glaucine, an aporphine alkaloid with anti-tussive, anti-inflammatory, and anti-nociceptive properties, has been identified in post-race samples from racehorses. To investigate pharmacokinetics of glaucine in horses, a three-way crossover study of intravenous and oral glaucine (0.1 mg/kg) and orally administered tulip poplar shavings (50 g shavings = 0.001 mg/kg glaucine) was performed in six horses. A two-compartment model best described IV administration with alpha ( t 1 / 2 α ) and beta ( t 1 / 2 ß ) half-life lives of 0.3 (0.1-0.7) and 3.1 (2.4-7.8) h, respectively. The area under the curve ( AUC 0 ∞ iv ) was 45.4 (34.7-52.3) h*ng/ml, and the volume of distribution of the central (Vdc ) and peripheral (Vdp ) compartments was 2.7 (1.3-4.6) and 4.9 (4.3-8.2) L/kg, respectively. A one compartment model best described the oral administration of glaucine with absorption ( t 1 / 2 ka ) and elimination ( t 1 / 2 kel ) half-lives of 0.09 (0.05-0.15) and 0.7 (0.6-0.8) h, respectively. The area under the curve ( AUC 0 ∞ PO ) was 15.1 (8.0-19.5) h·ng/ml. Bioavailability following oral administration was 17%-48%. Following ingestion of shavings, glaucine and liriodenine were detectable in plasma for up to 16 and 48 h, respectively. Glaucine was quantifiable briefly in the urine from two horses. Liriodenine was quantifiable in urine for 12-20 h in four horses and for 48 h in two horses. The presence of liriodenine indicates ingestion of tulip poplar tree parts, however, does not rule out co-administration of purified glaucine in horses.

Oxocrebanine from Stephania pierrei exerts macrophage anti-inflammatory effects by downregulating the NF-κB, MAPK, and PI3K/Akt signalling pathways.

Plant-derived medicinal compounds are increasingly being used to treat acute and chronic inflammatory diseases, which are generally caused by aberrant inflammatory responses. Stephania pierrei Diels, also known as Sabu-lueat in Thai, is a traditional medicinal plant that is used as a remedy for several inflammatory disorders. Since aporphine alkaloids isolated from S. pierrei tubers exhibit diverse pharmacological characteristics, we aimed to determine the anti-inflammatory effects of crude extracts and alkaloids isolated from S. pierrei tubers against lipopolysaccharide (LPS)-activated RAW264.7 macrophages. Notably, the n-hexane extract strongly suppressed nitric oxide (NO) while exhibiting reduced cytotoxicity. Among the five alkaloids isolated from the n-hexane extract, the aporphine alkaloid oxocrebanine exerted considerable anti-inflammatory effects by inhibiting NO secretion. Oxocrebanine also significantly suppressed prostaglandin E2, tumour necrosis factor-α, interleukin (IL)-1ß, IL-6, inducible nitric oxide synthase, and cyclooxygenase (COX)-2 protein expression by inactivating the nuclear factor κB, c-Jun NH2-terminal kinase, extracellular signal-regulated kinase 1/2, and phosphatidylinositol 3-kinase/Akt inflammatory signalling pathways. Molecular docking analysis further revealed that oxocrebanine has a higher affinity for toll-like receptor 4/myeloid differentiation primary response 88 signalling targets and the COX-2 protein than native ligands. Thus, our findings highlight the potential anti-inflammatory effects of oxocrebanine and suggest that certain alkaloids of S. pierrei could be used to treat inflammatory diseases.

A Systematic Method for the Identification of Aporphine Alkaloid Constituents in Sabia schumanniana Diels Using UHPLC-Q-Exactive Orbitrap/Mass Spectrometry

Sabia schumanniana Diels (SSD) is a plant whose stems are used in traditional folk medicine for the treatment of lumbago and arthralgia. Previous studies have revealed chemical constituents of SSD, including triterpenoids and aporphine alkaloids. Aporphine alkaloids contain a variety of active components, which might facilitate the effective treatment of lumbago and arthralgia. However, only 5-oxoaporphine (fuseine) has been discovered in SSD to date. In this study, we sought to systematically identify the aporphine alkaloids in SSD. We established a fast and reliable method for the detection and identification of these aporphine alkaloids based on ultra-high-performance liquid chromatography (UHPLC)-Q-Exactive-Orbitrap/mass spectrometry combined with parallel reaction monitoring (PRM). We separated all of the analyzed samples using a Thermo Scientific Hypersil GOLD™ aQ C18 column (100 mm × 2.1 mm, 1.9 µm). Finally, we identified a total of 70 compounds by using data such as retention times and diagnostic ions. No fewer than 69 of these SSD aporphine alkaloids have been reported here for the first time. These findings may assist in future studies concerning this plant and will ultimately contribute to the research and development of new drugs.

Natural potential neuroinflammatory inhibitors from Stephania epigaea H.S. Lo.

Stephania epigaea H. S. Lo is a folk medicine widely distributed in the south of China, especially in Yunnan and Guangxi province. An in vitro anti-neuroinflammatory study showed that total alkaloids of it can potently inhibit LPS-induced NO releasing of BV2 cells with an IC50 value of 10.05 ± 2.03 µg/mL (minocycline as the positive drug, IC50 15.49 ± 2.14 µM). The phytochemical investigation of the total alkaloids afforded three new phenanthrene (1-3), two lactams (4a, 4b), and nine aporphine derivatives (5-13). The final structure of 1 was identified by computer-assisted structure elucidation (ACD/Structure Elucidator software and the 13C NMR calculation with GIAO method) due to many possibilities of the substituent pattern. All isolates were evaluated for their anti-neuroinflammatory effects, and as a result, 5, 8, 10, and 11 exhibited stronger inhibitory activities than the minocycline. The results suggested S. epigaea could provide potential therapeutic agents for neurodegenerative diseases.

Aporphine Alkaloids from Ocotea puberula with Anti-Trypanosoma Cruzi Potential - Activity of Dicentrine-ß-N-Oxide in the Plasma Membrane Electric Potentials.

One new aporphine, dicentrine-ß-N-oxide (1), together with five related known alkaloids dehydrodicentrine (2), predicentrine (3), N-methyllaurotetanine (4), cassythicine (5), and dicentrine (6) were isolated from the leaves of Ocotea puberula (Lauraceae). Antiprotozoal activity of the isolated compounds was evaluated in vitro against trypomastigote forms of Trypanosoma cruzi. Among the tested compounds, alkaloid 1 exhibited higher potential with EC50 value of 18.2 µM and reduced toxicity against NCTC cells (CC50 >200 µM - SI>11.0), similar to positive control benznidazole (EC50 of 17.7 µM and SI=10.7). Considering the promising results of dicentrine-ß-N-oxide (1) against trypomastigotes, the mechanism of parasite death caused by this alkaloid was investigated. As observed, this compound reached the plasma membrane electric potential directly after 2 h of incubation and triggered mitochondrial depolarization, which probably leads to trypomastigote death. Therefore, dicentrine-ß-N-oxide (1), reported for the first time in this work, can contribute to future works for the development of new trypanocidal agents.