Biosensor and machine learning-aided engineering of an amaryllidaceae enzyme.

A major challenge to achieving industry-scale biomanufacturing of therapeutic alkaloids is the slow process of biocatalyst engineering. Amaryllidaceae alkaloids, such as the Alzheimer's medication galantamine, are complex plant secondary metabolites with recognized therapeutic value. Due to their difficult synthesis they are regularly sourced by extraction and purification from the low-yielding daffodil Narcissus pseudonarcissus. Here, we propose an efficient biosensor-machine learning technology stack for biocatalyst development, which we apply to engineer an Amaryllidaceae enzyme in Escherichia coli. Directed evolution is used to develop a highly sensitive (EC50 = 20 µM) and specific biosensor for the key Amaryllidaceae alkaloid branchpoint 4'-O-methylnorbelladine. A structure-based residual neural network (MutComputeX) is subsequently developed and used to generate activity-enriched variants of a plant methyltransferase, which are rapidly screened with the biosensor. Functional enzyme variants are identified that yield a 60% improvement in product titer, 2-fold higher catalytic activity, and 3-fold lower off-product regioisomer formation. A solved crystal structure elucidates the mechanism behind key beneficial mutations.

Narcissus' belief about his body: Aspects of narcissism, body image, and eating disorder symptoms.

OBJECTIVE: Narcissism may play a role in shaping body image concerns. Here we examined the relationships between narcissism (i.e., agentic extraversion, antagonism, narcissistic neuroticism, leadership/authority, exhibitionism/entitlement) and body image concerns and disturbances (i.e., drive for thinness, drive for muscularity, eating disorder symptoms, body mass index, current/desired fat, and current/desired muscularity). METHODS: Mechanical Turk workers from the USA (N = 430; 64% male) completed the Narcissistic Admiration and Rivalry Questionnaire, the Hypersensitive Narcissism Scale, the Narcissistic Personality Inventory, the Drive for Muscularity Scale, the Drive for Thinness Scale, the Eating Disorder Examination Questionnaire-Short, and the Sex-Specific Somatomorphic Matrixes. RESULTS: All narcissistic factors were associated with a greater drive for thinness (except for leadership/authority) and for muscularity, more eating disorder symptoms, a greater desired body fat (except for leadership/authority), and a greater current muscularity. Greater agentic extraversion and exhibitionism/entitlement were associated with lower levels of current body fat, and greater antagonism was associated with a greater desired muscularity. DISCUSSION: Notably, individual differences in narcissism appeared to be important in understanding body image concerns, broadly speaking. We found that narcissism may be associated with body image concerns among both sexes differently, and especially that drive for thinness was more related to narcissism in men. Our results emphasize the importance of narcissism in formulating and treating body image-related disorders for both men and women. Ultimately, narcissistic features of personality may be risk factors for developing and perpetuating body image concerns, and therefore should be considered in assessment, formulation, diagnosis, and treatment of eating disorders.

NtMYB1 and NtNCED1/2 control abscisic acid biosynthesis and tepal senescence in Chinese narcissus (Narcissus tazetta).

Chinese narcissus (Narcissus tazetta var. chinensis cv. 'Jinzhanyintai') is one of the 10 most famous traditional flowers of China, having a beautiful and highly ornamental flower with a rich fragrance. However, the flower longevity affects its commercial appeal. While petal senescence in Narcissus is ethylene-independent and abscisic acid-dependent, the regulatory mechanism has yet to be determined. In this study, we identified a R2R3-MYB gene (NtMYB1) from Narcissus tazetta and generated oeNtMYB1 and Ntmyb1 RNA interference mutants in Narcissus as well as an oeNtMYB1 construct in Arabidopsis. Overexpressing NtMYB1 in Narcissus or Arabidopsis led to premature leaf yellowing, an elevated level of total carotenoid, a reduced level of chlorophyll b, and a decrease in photosystem II fluorescence (Fv/Fm). A dual-luciferase assay and chromatin immunoprecipitation-quantitative PCR revealed that NtMYB1 directly binds to the promoter of NtNCED1 or NtNCED2 and activates NtNCED1/2 gene expression both in vitro and in vivo. Moreover, overexpressing NtMYB1 accelerated abscisic acid biosynthesis, up-regulated the content of zeatin and abscisic acid, and down-regulated the level of ß-carotene and gibberellin A1, leading to petal senescence and leaf yellowing in Narcissus. This study revealed a regulatory process that is fundamentally different between non-photosynthetic organs and leaves.

Advances on the Amaryllidacea Alkaloids Collected in South Africa, Andean South America and the Mediterranean Basin.

The alkaloids are one of the most represented family of natural occurring biological active compounds. Amaryllidaceae are also very well known for their beautiful flower and are thus used as ornamental plants in historic and public gardens. The Amaryllidacea alkaloids constitute an important group that is subdivided into different subfamilies with different carbon skeletons. They are well known from ancient times for their long application in folk medicine, and in particular, Narcissus poeticus L. was known to Hippocrates of Cos (ca. B.C. 460-370), who treated uterine tumors with a formulate prepared from narcissus oil. To date, more than 600 alkaloids of 15 chemical groups exhibiting various biological activities have been isolated from the Amaryllidaceae plants. This plant genus is diffused in regions of Southern Africa, Andean South America and the Mediterranean basin. Thus, this review describes the chemical and biological activity of the alkaloids collected in these regions in the last two decades as weel those of isocarbostyls isolated from Amaryllidaceae in the same regions and same period.

Under the thin skin of narcissus: Facial muscle activity reveals amplified emotional responses to negative social evaluation in individuals with grandiose narcissistic traits.

Individuals with grandiose narcissism exhibit enhanced antagonism and a defensive pattern of discordance between their emotional and physiological reactions to self-threatening evaluations. Although theoretical perspectives link narcissistic defensiveness to negative emotions, empirical evidence linking grandiose narcissism to emotional reactivity remains mixed. The current study used self-reported affect, electrocardiography, and facial electromyography (fEMG) to examine whether people scoring high in grandiose narcissism show amplified physiological and self-reported emotional reactivity to negative social evaluation. Following two challenging cognitive tasks, participants received negative and neutral feedback in a face-to-face evaluation situation. Receiving negative feedback decreased self-reported positive affect and dominance, slowed heart rate, and amplified fEMG activity related to frowning and eye constriction. Although self-reported emotional reactions were unrelated to grandiose narcissism, fEMG activity associated with negative affect was significantly enhanced by grandiose narcissism. In conclusion, individuals with higher levels of grandiose narcissism may not be willing to report overt emotional reactivity to self-threatening feedback, but physiological responses "beneath their thin skin" reveal amplified threat-related facial muscle activity suggestive of a negative emotional state.

Botanical Briefs: Daffodils (Narcissus Species).

Daffodils (Narcissus species) are the most common cause of irritant contact dermatitis among florists. Calcium oxalate crystals contained in the sap of the daffodil plants lead to irritant contact dermatitis on the skin. Daffodil rash commonly presents with fissuring, scaling, and erythema of the fingertips, hands, and forearms. The best preventative measure is to wear appropriate protective gloves and clothing.

Identification of haemanthamine as a phytotoxic alkaloid in Narcissus pseudonarcissus L. (Daffodil) emerging buds.

Emerging buds of Narcissus pseudonarcissus were found to accumulate the alkaloid haemanthamine (1) at high concentrations, exceeding that of narciclasine (2), the most abundant constituent in bulbs of the plant. A phytoactivity screening assay demonstrated the novel phytotoxicity of haemanthamine against Raphanus sativus (radish), Lactuca sativus (lettuce), Triticum aestivum (red wheat), Solanum lycopersicum (tomato), Cucumis sativus (cucumber), Ipomoea (Morning glory), and Lens culinaris (lentil). Haemanthamine (1) phytotoxicity was found to exceed that of the commercial herbicide glyphosate and less toxic than narciclasine (2).

Alkaloid Profile in Wild Autumn-Flowering Daffodils and Their Acetylcholinesterase Inhibitory Activity.

Amaryllidaceae alkaloids are secondary metabolites with interesting medicinal properties. Almost every Narcissus species can synthesize them and constitute an excellent source for their isolation and study. Several Amaryllidaceae alkaloids have shown acetylcholinesterase inhibitory activities and are a promising tool for treating cholinergic disorders such as Alzheimer's disease (AD). Indeed, three of the four palliative treatments approved for AD are acetylcholinesterase (AChE) inhibitors and one of them, galanthamine, is an Amaryllidaceae alkaloid itself. This molecule is currently isolated from natural sources. However, its production is insufficient to supply the increasing demand for the active principle. Our main aim is to discover tools to improve galanthamine production and to prospect for potential new and more efficient drugs for AD treatment. Furthermore, we seek to broaden the knowledge of plants of the genus Narcissus from a chemotaxonomic perspective. Hence, in this study, we evaluate the alkaloid content through GC-MS and the AChE inhibitory activity of ten autumn-flowering Narcissus, which have been less studied than their spring-flowering counterparts. A total of thirty Amaryllidaceae alkaloids have been found, twenty-eight properly identified. Two Narcissus contained galanthamine, and seven were able to inhibit AChE.

Synonymous Codon Usage Analysis of Three Narcissus Potyviruses.

Narcissus degeneration virus (NDV), narcissus late season yellows virus (NLSYV) and narcissus yellow stripe virus (NYSV), which belong to the genus Potyvirus of the family Potyviridae, cause significant losses in the ornamental value and quality of narcissus. Several previous studies have explored the genetic diversity and evolution rate of narcissus viruses, but the analysis of the synonymous codons of the narcissus viruses is still unclear. Herein, the coat protein (CP) of three viruses is used to analyze the viruses' phylogeny and codon usage pattern. Phylogenetic analysis showed that NYSV, NDV and NLSYV isolates were divided into five, three and five clusters, respectively, and these clusters seemed to reflect the geographic distribution. The effective number of codon (ENC) values indicated a weak codon usage bias in the CP coding region of the three narcissus viruses. ENC-plot and neutrality analysis showed that the codon usage bias of the three narcissus viruses is all mainly influenced by natural selection compared with the mutation pressure. The three narcissus viruses shared the same best optimal codon (CCA) and the synonymous codon prefers to use codons ending with A/U, compared to C/G. Our study shows the codon analysis of different viruses on the same host for the first time, which indicates the importance of the evolutionary-based design to control these viruses.

A novel plant lectin, NTL-125, interferes with SARS-CoV-2 interaction with hACE2.

COVID-19 caused by SARS-CoV-2 virus has had profound impact on the world in the past two years. Intense research is going on to find effective drugs to combat the disease. Over the past year several vaccines were approved for immunization. But SARS-CoV-2 being an RNA virus is continuously mutating to generate new variants, some of which develop features of immune escape. This raised serious doubts over the long-term efficacy of the vaccines. We have identified a unique mannose binding plant lectin from Narcissus tazetta bulb, NTL-125, which effectively inhibits SARS-CoV-2 replication in Vero-E6 cell line. In silico docking studies revealed that NTL-125 has strong affinity to viral Spike RBD protein, preventing it from attaching to hACE2 receptor, the gateway to cellular entry. Binding analyses revealed that all the mutant variants of Spike protein also have stronger affinity for NTL-125 than hACE2. The unique α-helical tail of NTL-125 plays most important role in binding to RBD of Spike. NTL-125 also interacts effectively with some glycan moieties of S-protein in addition to amino acid residues adding to the binding strength. Thus, NTL-125 is a highly potential antiviral compound of natural origin against SARS-CoV-2 and may serve as an important therapeutic for management of COVID-19.

Pressurized Natural Deep Eutectic Solvent Extraction of Galanthamine and Related Alkaloids from Narcissus pseudonarcissus.

The isolation of a compound from a natural source involves many organic and mostly toxic solvents for extraction and purification. Natural deep eutectic solvents have been shown to be efficient options for the extraction of natural products. They have the advantage of being composed of abundantly available common primary metabolites, being nontoxic and environmentally safe solvents. The aim of this study was to develop a natural deep eutectic solvent-based extraction method for galanthamine, an important therapeutic agent for the treatment of Alzheimer's disease. This alkaloid can be produced by synthesis or by extraction from Narcissus bulbs. To develop an efficient extraction method, a number of different natural deep eutectic solvents was first tested for their solubilization capacity of galanthamine bromide salt. Promising results were obtained for ionic liquids, as well as some amphoteric and acidic natural deep eutectic solvents. In a two-cycle extraction process, the best solvents were tested for the extraction of galanthamine from bulbs. The ionic liquids produced poor yields, and the best results were obtained with some acid and sugar mixtures, among which malic acid-sucrose-water (1 : 1 : 5) proved to be the best, showing similar yields to that of the exhaustive Soxhlet extraction with methanol. Furthermore, the natural deep eutectic solvent was more selective for galanthamine.

Narcissus Plants: A Melting Pot of Potyviruses.

Our paper presents detailed evolutionary analyses of narcissus viruses from wild and domesticated Narcissus plants in Japan. Narcissus late season yellows virus (NLSYV) and narcissus degeneration virus (NDV) are major viruses of Narcissus plants, causing serious disease outbreaks in Japan. In this study, we collected Narcissus plants showing mosaic or striped leaves along with asymptomatic plants in Japan for evolutionary analyses. Our findings show that (1) NLSYV is widely distributed, whereas the distribution of NDV is limited to the southwest parts of Japan; (2) the genomes of NLSYV isolates share nucleotide identities of around 82%, whereas those of NDV isolates are around 94%; (3) three novel recombination type patterns were found in NLSYV; (4) NLSYV comprises at least five distinct phylogenetic groups whereas NDV has two; and (5) infection with narcissus viruses often occur as co-infection with different viruses, different isolates of the same virus, and in the presence of quasispecies (mutant clouds) of the same virus in nature. Therefore, the wild and domesticated Narcissus plants in Japan are somewhat like a melting pot of potyviruses and other viruses.

Complete genome sequence of vallota mosaic virus detected in a narcissus bulb imported from the United States to Japan.

Narcissus (Narcissus albidus) imported from the United States exhibited leaf chlorosis during post-entry quarantine. We employed next-generation sequencing (NGS) on symptomatic leaf samples and detected vallota mosaic virus (ValMV), belonging to the genus Potyvirus, family Potyviridae, as the viral agent. Sanger sequencing of PCR products and rapid amplification of cDNA ends based on NGS contigs revealed that ValMV is 9,451 nucleotides (nt) in length, excluding the poly(A) tail. Nucleotide and amino acid (aa) sequences of the coat protein region had over 98% identity to previously reported ValMV isolates. In each of the 10 regions encoding mature proteins, however, the sequence identity to other potyviruses was 49.5-71.9% nt and 18.3-78.9% aa, values that are below the species demarcation thresholds for the family Potyviridae. Phylogenetic analysis revealed that our ValMV isolate is most closely related to known ValMV isolates and is grouped with other potyviruses. Taken together, our results indicate that the newly isolated ValMV belongs to a distinct species in the genus Potyvirus. This study provides the first report of the complete ValMV genome sequence and the first record of this virus in narcissus.

Supercritical CO2 Extraction of Narcissus poeticus L. Flowers for the Isolation of Volatile Fragrance Compounds.

The flowers of Narcissus poeticus are used for the isolation of valuable fragrance substances. So far, as the majority of these substances consist of volatile and sensitive to heat compounds, there is a need of developing effective methods for their recovery. In this study, freeze-dried N. poeticus inflorescences were extracted with pure supercritical CO2 (SFE-CO2) and its mixture with 5% co-solvent ethanol (EtOH) at 40 °C. Extract yields varied from 1.63% (12 MPa) to 3.12% (48 MPa, 5% EtOH). In total, 116 volatile compounds were identified by GC-TOF/MS in the extracts, which were divided into 20 different groups. Benzyl benzoate (9.44-10.22%), benzyl linoleate (1.72-2.17%) and benzyl alcohol (0.18-1.00%) were the major volatiles among aromatic compounds. The amount of the recovered benzyl benzoate in N. poeticus SFE-CO2 extracts varied from 58.98 ± 2.61 (24 MPa) to 91.52 ± 1.36 (48 MPa) mg/kg plant dry weight (pdw). α-Terpineol dominated among oxygenated monoterpenes (1.08-3.42%); its yield was from 9.25 ± 0.63 (12 MPa) to 29.88 ± 1.25 (48 MPa/EtOH) mg/kg pdw. Limonene was the major monoterpene hydrocarbon; (3E)-hexenol and heneicosanol dominated among alcohols and phenols; dihydroactinidiolide and 4,8,12,16-tetramethyl heptadecan-4-olide were the most abundant lactones; heptanal, nonanal, (2E,4E)-decadienal and octadecanal were the most abundant aldehydes. The most important prenol lipids were triterpenoid squalene, from 0.86 ± 0.10 (24 MPa) to 7.73 ± 0.18 (48 MPa/EtOH) mg/kg pdw and D-α-tocopherol, from 1.20 ± 0.04 (12 MPa) to 15.39 ± 0.31 (48 MPa/EtOH) mg/kg pdw. Aliphatic hydrocarbons (waxes) constituted the main part (41.47 to 54.93%) in the extracts; while in case of a 5% EtOH the percentage of alkanes was the lowest. The fraction of waxes may be removed for the separation of higher value fragrance materials. In general, the results obtained are promising for a wider application of SFE-CO2 for the recovery of fragrance substances from N. poeticus flowers.

Development of a protocol for the elimination of Cyrtanthus elatus virus-A from Narcissus tazetta by in vitro chemotherapy in combination with electrotherapy.

Narcissus (Narcissus tazetta) is a bulbous ornamental plant propagated vegetatively from bulbs. The Cyrtanthus elatus virus-A (CyEV-A) had been reported to cause a severe mosaic and yellow stripe disease in narcissus. Therefore, this study aimed to develop a protocol for the elimination of CyEV-A from infected bulblets by in vitro chemotherapy (30-50 mg/L ribavirin for 30 days) and electrotherapy (10-30 mA for 20 min), individually and in combination, to produce virus-free plants. The regenerated plants obtained from these treatments were screened for the absence of the CyEV-A by reverse-transcription polymerase chain reaction assays using a set of degenerate primers specific for a potyvirus coat protein gene. The results showed that in vitro chemotherapy (30 mg/L ribavirin for 30 days) alone produced 46.0 % (14/30) of virus-free plants, while electrotherapy (20 mA for 20 min) alone produced 40.0 % (12/30) of virus-free plants. In comparison, a combination of chemotherapy (30 mg/L ribavirin for 30 days) and electrotherapy (20 mA for 20 min) produced 50.0 % (15/30) of virus-free plants. The virus-free plants obtained from this combination treatment exhibited better growth and produced more bulbs compared to the other treatments and control. The protocol may be used for the control of the virus disease in narcissus.

Transcriptome-Based WGCNA Analysis Reveals Regulated Metabolite Fluxes between Floral Color and Scent in Narcissus tazetta Flower.

A link between the scent and color of Narcissus tazetta flowers can be anticipated due to their biochemical origin, as well as their similar biological role. Despite the obvious aesthetic and ecological significance of these colorful and fragrant components of the flowers and the molecular profiles of their pigments, fragrant formation has addressed in some cases. However, the regulatory mechanism of the correlation of fragrant components and color patterns is less clear. We simultaneously used one way to address how floral color and fragrant formation in different tissues are generated during the development of an individual plant by transcriptome-based weighted gene co-expression network analysis (WGCNA). A spatiotemporal pattern variation of flavonols/carotenoids/chlorophyll pigmentation and benzenoid/phenylpropanoid/ monoterpene fragrant components between the tepal and corona in the flower tissues of Narcissus tazetta, was exhibited. Several candidate transcription factors: MYB12, MYB1, AP2-ERF, bZIP, NAC, MYB, C2C2, C2H2 and GRAS are shown to be associated with metabolite flux, the phenylpropanoid pathway to the production of flavonols/anthocyanin, as well as related to one branch of the phenylpropanoid pathway to the benzenoid/phenylpropanoid component in the tepal and the metabolite flux between the monoterpene and carotenoids biosynthesis pathway in coronas. It indicates that potential competition exists between floral pigment and floral fragrance during Narcissus tazetta individual plant development and evolutionary development.

Potential antioxidant effects of Narcissus tazetta phenolic compounds against cadmium chloride-induced hepatotoxicity in Swiss albino mice.

Narcissus tazetta (Amaryllidaceae) is a medicinal plant widely used for cut flowers and potted ornamental plant in Tunisia flora. The current study evaluated the phenolic composition and antioxidant properties of its flower extracts and investigated its potential protective activity against cadmium chloride (CdCl2)-induced hepatotoxicity in mice. Mice were divided into six groups of six each: group 1, serving as negative controls, received by intraperitoneal way only distilled water; group 2 received by intraperitoneal way CdCl2 (0.16 mg/kg bw); groups 3 and 4 received CdCl2 at the same dose of group 2 and 100 or 200 mg/kg bw of Narcissus tazetta flower extracts via oral route; groups 5 and 6, serving as positive controls, received only Narcissus tazetta flower extracts. Polyphenolic compounds of the extract were analyzed by colorimetric and high-performance liquid chromatography-mass spectrometry (HPLC-MS) methods. Total antioxidant activity and 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging potential of the extract were estimated using colorimetric method. Results indicated that ethanolic flower extract contained high levels of total phenolic and flavonoid along with a strong total antioxidant and DPPH free radical scavenging activities. HPLC-MS analysis identified eight phenolic compounds, including rutin, kaempferol glycosides, and chlorogenic acids. The extract also exhibited marked hepatoprotective effects against CdCl2 toxicity by reducing hepatic levels of malondialdehyde, advanced oxidation protein products, hydrogen peroxide, metallothioneins, and DNA degradation. Additionally, co-administration of Narcissus tazetta flower extracts lowered the plasma activities of transaminases, gamma glutamyl transpeptidase, and lactate dehydrogenase and increased hepatic levels of reduced glutathione, nonprotein thiols, vitamin C, and catalase activity. The hepatoprotective effects of the extract were demonstrated by histopathological improvement of liver disorders. The current study provided ethnopharmacological application of Narcissus tazetta flower extracts against CdCl2-induced oxidative stress, suggesting its chemoprevention role of its phenolic compounds as a natural antioxidant.

NtbHLH1, a JAF13-like bHLH, interacts with NtMYB6 to enhance proanthocyanidin accumulation in Chinese Narcissus.

BACKGROUND: Flavonoid biosynthesis in plants is primarily regulated at the transcriptional level by transcription factors modulating the expression of genes encoding enzymes in the flavonoid pathway. One of the most studied transcription factor complexes involved in this regulation consists of a MYB, bHLH and WD40. However, in Chinese Narcissus (Narcissus tazetta L. var. chinensis), a popular monocot bulb flower, the regulatory mechanism of flavonoid biosynthesis remains unclear. RESULTS: In this work, genes related to the regulatory complex, NtbHLH1 and a R2R3-MYB NtMYB6, were cloned from Chinese Narcissus. Phylogenetic analysis indicated that NtbHLH1 belongs to the JAF13 clade of bHLH IIIf subgroup, while NtMYB6 was highly homologous to positive regulators of proanthocyanidin biosynthesis. Both NtbHLH1 and NtMYB6 have highest expression levels in basal plates of Narcissus, where there is an accumulation of proanthocyanidin. Ectopic over expression of NtbHLH1 in tobacco resulted in an increase in anthocyanin accumulation in flowers, and an up-regulation of expression of the endogenous tobacco bHLH AN1 and flavonoid biosynthesis genes. In contrast, the expression level of LAR gene was significantly increased in NtMYB6-transgenic tobacco. Dual luciferase assays showed that co-infiltration of NtbHLH1 and NtMYB6 significantly activated the promoter of Chinese Narcissus DFR gene. Furthermore, a yeast two-hybrid assay confirmed that NtbHLH1 interacts with NtMYB6. CONCLUSIONS: Our results suggest that NtbHLH1 may function as a regulatory partner by interacting directly with NtMYB6 to enhance proanthocyanidin accumulation in Chinese Narcissus.

GC-MS analysis of Amaryllidaceae and Sceletium-type alkaloids in bioactive fractions from Narcissus cv. Hawera.

RATIONALE: Narcissus cv. Hawera has been found to biosynthesize some Sceletium-type alkaloids with antidepressant and anxiolytic activities. This ornamental plant has been poorly studied as a source of bioactive alkaloids including some contraversive reports on in vitro and intact plants. In this study, a detailed GC-MS characterization of its alkaloid fractions is presented. METHODS: GC-MS was used for the identification of compounds in the alkaloid fractions. Both underivatized and silylated samples were analyzed simultaneously. Elevated plus maze and tail suspension tests were used to assay the anxiolytic and antidepressant activities. Ellman's and MTT-dye reduction assays were used to evaluate the acetylcholinesterase (AChE) inhibitory and cytotoxicity activities, respectively. RESULTS: Of the 29 alkaloids, 13 of Sceletium-type were detected. Two new alkaloids were identified as 2-oxo-mesembrine and 2-oxo-epi-mesembrenol. Lycorine was found as a major compound (43.5%) in the crude silylated methanol extract. After the elimination of lycorine by pre-crystallization, the major alkaloids were 40.8% 6-epi-mesembranol, 16.2% 6-epi-mesembrenol, and 13.8% sanguinine. This fraction showed anxiolytic and antidepressant-like activities as well as potent AChE inhibitory and antineoplastic activities. CONCLUSIONS: Silylation of the alkaloid fractions from Narcissus cv. Hawera provides better separation, structural information, and improved sensitivity for compounds with two and more hydroxyl groups. The lycorine-free alkaloid fraction shows a great potential for further pharmacological studies.

Integrating Narcissus-derived galanthamine production into traditional upland farming systems.

Alzheimer's disease (AD) is a disorder associated with progressive degeneration of memory and cognitive function. Galantamine is a licenced treatment for AD but supplies of the plant alkaloid that it is produced from, galanthamine, are limited. This three-year system study tested the potential to combine Narcissus-derived galanthamine production with grassland-based ruminant production. Replicate plots of permanent pasture were prepared with and without bulbs of Narcissus pseudonarcissus sown as lines into the sward. Two different fertiliser regimes were imposed. The above-ground green biomass of N. pseudonarcissus was harvested in early spring and the galanthamine yield determined. In the second harvest year a split-plot design was implemented with lines of N. pseudonarcissus cut annually and biennially. All plots were subsequently grazed by ewes and lambs and animal performance recorded. Incorporation of N. pseudonarcissus into grazed permanent pasture had no detrimental effects on the health or performance of the sheep which subsequently grazed the pasture. There was no consistency to the effects of fertiliser rates on galanthamine yields. There was no difference in overall galanthamine yield if N. pseudonarcissus was cut biennially (1.64 vs. 1.75 kg galanthamine/ha for annual combined vs biennial cuts respectively; s.e.d = 0.117 kg galanthamine/ha; ns). This study verified the feasibility of a dual cropping approach to producing plant-derived galanthamine.