Alkaloid Profiles Research Articles

Metabolite Fingerprinting of Piper nigrum L. from Different Regions of Thailand by UHPLC-QTOF-MS Approach and In Vitro Bioactivities

This study aimed to profile the metabolites, piperine content, and biological activities (antioxidant, anti-inflammatory, anticancer, antibacterial, and apoptotic cell) of 15 samples of Piper nigrum L. from various geographic regions of Thailand. UHPLC-QTOF-MS was applied to establish alkaloid profiles of the P. nigrum L. samples. PCA and HCA analysis showed that the samples from the PN-1, PN-2, and PN-11 sources differed significantly from those of other sources in terms of compound type and alkaloid concentration profile. Comparative analysis based on a validated HPLC-DAD method was used for the standardization of 15 varieties of P. nigrum L. with piperine as a standard phytomarker which revealed that PN-4 source possessed a high content of piperine. PN-2 showed the highest antioxidant capacity (DPPH activity) while PN-10, PN-12, and PN-15 showed the highest FRAP activity. PN-11 source showed high inhibition of anti-inflammatory and anticancer properties. Furthermore, PN-3, and PN-4 also showed the highest both anti-inflammatory and antibacterial activity. Almost samples have been shown to induce late apoptosis and necrosis in SW480 cell line. This study will also be useful for the development of quality control parameters through the standardization of varieties of P. nigrum L. as raw materials in traditional medicine formulations. HIGHLIGHTS Metabolite profiling of the nigrum L. varieties from 15 different geographical locations in Thailand Quality control parameter for standardization based on marker analysis by HPLC-DAD using piperine as the marker compound Comparisons of metabolites among 15 different cultivations by Principal Component Analysis (PCA) and hierarchical cluster analysis (HCA) Biological activity of the nigrum L. varieties from 15 different geographical locations in Thailand GRAPHICAL ABSTRACT

Transcriptome and Metabolome Combined to Analyze Quinoa Grain Quality Differences of Different Colors Cultivars.

Quinoa (Chenopodium quinoa Wild.) has attracted considerable attention owing to its unique nutritional, economic, and medicinal values. Meanwhile, quinoa germplasm resources and grain colors are rich and diverse. In this study, we analyzed the composition of primary and secondary metabolites and the content of the grains of four different high-yield quinoa cultivars (black, red, white, and yellow) harvested 42 days after flowering. The grains were subjected to ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) and transcriptome sequencing to identify the differentially expressed genes and metabolites. Analysis of candidate genes regulating the metabolic differences among cultivars found that the metabolite profiles differed between white and black quinoa, and that there were also clear differences between red and yellow quinoa. It also revealed significantly altered amino acid, alkaloid, tannin, phenolic acid, and lipid profiles among the four quinoa cultivars. Six common enrichment pathways, including phenylpropane biosynthesis, amino acid biosynthesis, and ABC transporter, were common to metabolites and genes. Moreover, we identified key genes highly correlated with specific metabolites and clarified the relationship between them. Our results provide theoretical and practical references for breeding novel quinoa cultivars with superior quality, yield, and stress tolerance. Furthermore, these findings introduce an original approach of integrating genomics and transcriptomics for screening target genes that regulate the desirable traits of quinoa grain.

Effects of Heating, Pelleting, and Feed Matrix on Apparent Concentrations of Cereal Ergot Alkaloids in Relation to Growth Performance and Welfare Parameters of Backgrounding Beef Steers.

As the contamination of cereal grains with ergot has been increasing in Western Canada, studies were undertaken to evaluate the impacts of heating (60, 80, 120, or 190 °C) alone or in combination with pelleting on concentrations of ergot alkaloids. Fifteen samples of ergot-contaminated grain from Alberta and Saskatchewan were assayed for R and S epimers of six alkaloids (ergocryptine, ergocristine, ergocornine, ergometrine, ergosine, and ergotamine) using HPLC MS/MS. Five samples with distinct alkaloid profiles were then selected for heating and pelleting studies. Heating resulted in a linear increase (p < 0.05) of total R and total S epimers with increasing temperature, although some individual R epimers were stable (ergometrine, ergosine, ergotamine). Pelleting also increased (p < 0.05) concentrations of total R and total S epimers detected, although ergometrine concentration decreased (p < 0.05) after pelleting. A feeding study arranged in a 2 × 2 factorial structure used 48 backgrounding Angus-cross steers fed four different diets: (1) Control Mash (CM, no added ergot), (2) Control Pellet (CP), (3) Ergot Mash (EM), or (4) Ergot Pellet (EP). Pelleting heated the ergot to 90–100 °C under 4 bars pressure, but the ergot used in the feeding study was not otherwise heated. Alkaloid concentrations of EM and EP varied by up to 1.1 mg/kg depending on the feed matrix assayed. No differences among treatments were noted for growth performance, feed intake, feed conversion, concentrations of serum prolactin and haptoglobin, hair cortisol, or in temperatures of extremities measured by infrared thermography. The only negative impacts of ergot alkaloids were on blood parameters indicative of reduced immune function or chronic inflammation. Pelleting did not heighten the negative clinical outcomes of ergot, although alkaloid concentrations of pelleted feed increased depending on the matrix assayed. It was hypothesized that the heat and pressure associated with pelleting may enhance the recovery of alkaloids from pelleted feed.

What Goes in Must Come Out? The Metabolic Profile of Plants and Caterpillars, Frass, And Adults of Asota (Erebidae: Aganainae) Feeding on Ficus (Moraceae) in New Guinea.

Insect herbivores have evolved a broad spectrum of adaptations in response to the diversity of chemical defences employed by plants. Here we focus on two species of New Guinean Asota and determine how these specialist moths deal with the leaf alkaloids of their fig (Ficus)hosts. As each focal Asota species is restricted to one of three chemically distinct species of Ficus, we also test whether these specialized interactions lead to similar alkaloid profiles in both Asota species. We reared Asota caterpillars on their respective Ficus hosts in natural conditions and analyzed the alkaloid profiles of leaf, frass, caterpillar, and adult moth samples using UHPLC-MS/MS analyses. We identified 43 alkaloids in our samples. Leaf alkaloids showed various fates. Some were excreted in frass or found in caterpillars and adult moths. We also found two apparently novel indole alkaloids-likely synthesized de novo by the moths or their microbiota-in both caterpillar and adult tissue but not in leaves or frass. Overall, alkaloids unique or largely restricted to insect tissue were shared across moth species despite feeding on different hosts. This indicates that a limited number of plant compounds have a direct ecological function that is conserved among the studied species. Our results provide evidence for the importance of phytochemistry and metabolic strategies in the formation of plant-insect interactions and food webs in general. Furthermore, we provide a new potential example of insects acquiring chemicals for their benefit in an ecologically relevant insect genus.

Indole-diterpenes alkaloid profiles of native grasses involved in tremorgenic syndromes in the Argentine Patagonia

One of the main intoxications to livestock in the Patagonia region of Argentina is the tremorgenic disease “Mal de Huecú", attributed to the consumption of the native grasses Poa huecu and/or Festuca argentina. In this report, five outbreaks of spontaneous intoxications were investigated. Several indole-diterpene alkaloids were identified in Poa huecu and Festuca argentina including the known tremorgen terpendole C and are likely the cause of “Mal de Huecú" disease.

Chemical composition of Ayahuasca seized in Poland

Ayahuasca is a traditional psychoactive brew of Amazonian indigenous, used for medicinal and spiritual purposes. Over the years, the consumption of ayahuasca all over the world has increased not only in shamanic and religious rituals, but also for recreational purposes. Wide variation in the concentration of alkaloids, i.e. N,N-dimethyltryptamine (DMT), harmaline, harmine, tetrahydroharmine (THH) and harmalol in Ayahuasca has been reported worldwide. Being a plant preparation, traditional Ayahuasca is variable in appearance, taste, and effects. Several studies suggest that it has anxiolytic, antidepressant, and antiaddictive effects. The aim of our study was to determine alkaloids concentration in Ayahuasca preparation seized in two cases of Ayahuasca rituals in Poland. In these cases we also analyzed plant material, i.e. the bark and stem of Banisteriopsis caapi, leaves of Psychotria viridis, seeds of Peganum Harmala and brown powder substances. The research material consisted of 32 samples of Ayahuasca brew seized in two Ayahuasca rituals cases in Poland and sent for examination to the Institute of Forensic Research in Krakow. Samples were prepared by dilution in ethanol or ethanol:water (1:1 v/v) mixture. The qualitative analyses of the samples were carried out by gas chromatography coupled with mass spectrometry (GC-MS). Gas chromatograph HP 6890N GC System coupled with the 5973 Network Mass Selective Detector with a quadrupole mass analyzer from Agilent Technologies (USA), were used in the study. Chromatographic separation of the tested compounds in the programmed temperature increase was carried out on the HP-5MS capillary column (30 m × 0.25 mm; 0.25 μm). The sample was dosed automatically in the splitless mode. The mass spectrometer operated in positive electron ionisation mode (EJ), and the energy beam was 70 eV. The acquisition was conducted in the scanning mode of the full mass range of m/z from 29 to 600 amu. The concentrations of DMT, harmaline and harmine were determined by liquid chromatography combined with mass spectrometry, a hybrid combination of quadrupole and time of flight analyzer (LC-MS/QTOF). The 1200 series liquid chromatograph and the 6520 Accurate-Mass QTOFMS mass spectrometer from Agilent Technologies (Santa Clara, CA, USA) were used. The separation was carried out on the Ascentis express C18 column (7.5 cm × 2.1 mm; 2.7 μm) from Supelco, thermostated at 35 °C. A mixture of 0.1% formic acid in water (A, v/v) and acetonitrile (B, v/v) was used as a mobile phase. Our research showed large variations in alkaloid profiles of Ayahuasca samples from one source. The concentration ranges of alkaloids was within: for DMT: 0.09–1.60 mg/mL, harmaline: 0.02–5.39 mg/mL and for harmine: 0.11–1.49 mg/mL. Concentration of DMT, HRL and HRM in Ayahuasca preparation was analyzed. Our research showed large variations in alkaloid profiles of Ayahuasca samples from one source. Variability of the brew composition and individual variability of effects together make it difficult for users to assess the quality of the brew that they drink during the ceremony. Our research results may be surely used as warning for participants of illicit Ayahuasca ceremonials, by increasing awareness about the quality of herbal preparations used.

Noninvasive Detection of Chemical Defenses in Poison Frogs Using the MasSpec Pen.

Poison frogs are well-known for their fascinating ability to store alkaloids in their skin as chemical defense against predators. Chemical methods used to study these alkaloids are limited by requirements for euthanasia or stress during sampling. Here, we demonstrate sensitive and biocompatible alkaloid detection and monitoring in vivo using the MasSpec Pen, a handheld, noninvasive chemical detection device coupled to a mass spectrometer. The MasSpec Pen allowed rapid (<15 s), gentle, and consecutive molecular analysis without harm or undue stress to the animals. Through a month-long alkaloid-feeding study with the dyeing poison frog, we observed temporal dynamics of chemical sequestration in vivo by comparing frogs fed the alkaloid decahydroquinoline (DHQ) to vehicle-fed frogs. We also demonstrate the feasibility of the MasSpec Pen for the untargeted detection of rich alkaloid profiles from skin extracts of the Diablito poison frog, collected from two distinct geographical populations in Ecuador. The results obtained in this study demonstrate the utility of the MasSpec Pen for direct, rapid, and biocompatible analysis of poison frog alkaloids.

Alkaloid Composition and Biological Activities of the Amaryllidaceae Species Ismene amancaes (Ker Gawl.) Herb.

Natural products have always played a significant role in the search for new drugs. One of the most relevant alkaloid-containing plant groups is the Amaryllidaceae family, a source of exclusive structures with a wide variety of pharmacological activities. The aim of this work was to determine the alkaloid composition and biological potential of an extract from the bulbs of an endemic Peruvian Amaryllidaceae species Ismene amancaes (Ker Gawl.) Herb. The alkaloid profiling was carried out by GC-MS, which revealed the presence of 13 compounds, 2 of them unidentified. The plant extract was found to contain high amounts of lycoramine, a galanthamine-type alkaloid. The extract also presented low inhibitory potential against the enzymes AChE and BuChE, with IC50 values of 14.6 ± 0.6 and 37.6 ± 1.4 μg·mL−1, respectively, and good to moderate inhibitory activity against the protozoan Plasmodium falciparum strain FCR-3 (chloroquine-resistant), with IC50 values of 3.78 ± 0.3 μg·mL−1. This is the first report of the alkaloid profile of a plant of the Ismene genus, which could be an interesting source of bioactive compounds.

Assessment of Tissue Specific Distribution and Seasonal Variation of Alkaloids in Alstonia scholaris.

Alstonia scholaris is a well-known source of alkaloids and widely recognized for therapeutic purposes to treat the ailments in human and livestock. However, the composition and production of alkaloids vary due to tissue specific metabolism and seasonal variation. This study investigated alkaloids in leaves, stems, trunk barks, fruits, and flowers of A. scholaris. The impact of seasonal changes on the production of alkaloids in the leaves of A. scholaris was also investigated. One and two-dimensional Nuclear Magnetic Resonance (NMR) experiments were utilized for the characterization of alkaloids and total eight alkaloids (picrinine, picralinal, akuammidine, 19 S scholaricine, 19,20 E vallesamine, Nb-demethylalstogustine N-Oxide, Nb-demethylalstogustine, and echitamine) were characterized and quantified. Quantitative and multivariate analysis suggested that the alkaloids content is tissue specific, illustrating the effect of plant tissue organization on alkaloidal production in A. scholaris. The results suggest that the best part to obtain alkaloids is trunk barks, since it contains 7 alkaloids. However, the best part for isolating picrinine, picralinal, akuammidine, 19 S scholaricine, and 19,20 E vallesamine is fruit, since it shows highest amount of these alkaloids. Undoubtedly, NMR and statistical methods are very helpful to differentiate the profile of alkaloids in A. scholaris.

The 168-year taxonomy of Claviceps in the light of variations: From three morphological species to four sections based on multigene phylogenies

Ergot fungi produce toxic sclerotia that have significant impacts on the agricultural, food and pharmaceutical industries. These fungi were classified in various genera (such as Sclerotium clavus, or Spermoedia clavus) before Tulasne, in 1853, erected Claviceps and described three species based on variations in morphology and host ranges, viz. C. purpurea, C. microcephala, and C. nigricans. Since then, knowledge regarding the biological, clinical, and pharmaceutical perspectives of ergot has accumulated rapidly. However, a serious taxonomic examination was lacking until Langdon’s revision accepted 25 species in 1952. That was followed by intensive regional studies by Loveless in the 1960s for Rhodesia (now Zimbabwe, Africa), and Tanda in Japan (1970s–1990s). More species names were reported and currently over 90 named taxa (species, varieties) are recorded in fungal name repositories (Mycobank and Index Fungorum). Most species were described based on morphological characteristics (sclerotia, ascomata and conidia) and host ranges. Some were characterized by their alkaloid profiles. Recently, DNA multi-locus sequence analyses (MLSA) were applied to resolve species complexes. For instance, the C. purpurea complex was separated into four species, and additional new species were recognized from South Africa and Canada. Infra- and supra-specific level genetic variations were identified via multi-locus and genomic studies. Based on five-locus phylogenies, Píchová and colleagues separated Claviceps into four sections: C. sect. Claviceps, C. sect. Citrinae, C. sect. Paspalorum and C. sect. Pusillae, for 60 species. Among these sections, several doubtful species names require clarification. Careful research on type specimens combined with molecular analyses is essential for clarifying these names.

Azotobacter chroococcum and Pseudomonas putida enhance pyrroloquinazoline alkaloids accumulation in Adhatoda vasica hairy roots by biotization

Adhatoda vasica is used in the treatment of cold, cough, chronic bronchitis, asthma, diarrhea, and dysentery. The biological activities of this species are attributed with the presence of alkaloids, triterpenoids, and flavonoids. Agrobacterium rhizogenes-mediated transformation of A. vasica, produces pyrroloquinazoline alkaloids, was achieved by infecting leaf discs with strain ATCC15834. The bacterial strain infected 82.7% leaf discs and 5–7 hairy root initials were developed from the cut edges of leaf discs. In this study, seven strains of Azotobacter chroococcum and five strains of Pseudomonas putida were used for the biotization of hairy roots. Plant growth-promoting rhizobacteria (PGPR) develops symbiotic association with roots of plants and increases the growth parameters of plants. PGPR (A. chroococcum and P. putida) increased the profiles of nitrogenase and acid phosphatase enzymes, biomass, dry matter contents, anthranilate synthase activity and accumulation of pyrroloquizoline alkaloids in the biotized hairy roots. Both enzymes (nitrogenase and acid phosphatase) maintain sufficient supply of nitrogen and dissolved phosphorus to the cells of hairy roots therefore, the levels of anthranilate synthase activity and pyrroloquinazoline alkaloids are increased. Total seven pyrroloquinazoline alkaloids (vasicine, vasicinone, vasicine acetate, 2-acetyl benzyl amine, vasicinolone, deoxyvasicine and vasicol) were identified from the biotized hairy roots of A. vasica. In our study, biotization increased the profiles of pyrroloquinazoline alkaloids therefore, this strategy may be used in increasing the production of medicinally important secondary metabolites in other plant species also. Our hypothetical model demonstrates that P. putida cell surface receptors receive root exudates by attaching on hairy roots. After attachment, the bacterial strain penetrates in the biotized hairy roots. This endophytic interaction stimulates acid phosphatase activity in the cells of biotized hairy roots. The P. putida plasmid gene (ppp1) expression led to the synthesis of acid phosphatase in cytosol. The enzyme enhances phosphorus availability as well as induces the formation of phosphoribosyl diphosphate. Later, phosphoribosyl diphosphate metabolizes to tryptophan and finally tryptophan converts to anthranilic acid. The synthesized anthranilic acid used in the synthesis of alkaloids in A. vasica.

Kratom (Mitragyna speciosa) Validation: Quantitative Analysis of Indole and Oxindole Alkaloids Reveals Chemotypes of Plants and Products.

Many consumers are turning to kratom ( Mitragyna speciosa ) to self-manage pain and opioid addiction. In the United States, an array of capsules, powders, and loose-leaf kratom products are readily available. Additionally, several online sites supply live kratom plants. A prerequisite to establishing quality control and quality assurance standards for the kratom industry, or understanding how alkaloid levels effect clinical outcomes, is the identification and quantitation of major and minor alkaloid constituents within available products and preparations. To this end, an ultra-high performance liquid chromatography-high resolution mass spectrometry method was developed for the analysis of 8 indole alkaloids (7-hydroxymitragynine, ajmalicine, paynantheine, mitragynine, speciogynine, isopaynantheine, speciociliatine, and mitraciliatine) and 6 oxindole alkaloids (isomitraphylline, isospeciofoleine, speciofoline, corynoxine A, corynoxeine, and rhynchophylline) in US-grown kratom plants and commercial products. These commercial products shared a qualitatively similar alkaloid profile, with 12 – 13 detected alkaloids and high levels of the indole alkaloid mitragynine (13.9 ± 1.1 – 270 ± 24 mg/g). The levels of the other major alkaloids (paynantheine, speciociliatine, speciogynine, mitraciliatine, and isopaynantheine) and the minor alkaloids varied in concentration from product to product. The alkaloid profile of US-grown M. speciosa “Rifat” showed high levels of the indole alkaloid speciogynine (7.94 ± 0.83 – 11.55 ± 0.18 mg/g) and quantifiable levels of isomitraphylline (0.943 ± 0.033 – 1.47 ± 0.18 mg/g). Notably, the alkaloid profile of a US-grown M. speciosa seedling was comparable to the commercial products with a high level of mitragynine (15.01 ± 0.20 mg/g). This work suggests that there are several M. speciosa chemotypes.

Anti-Trypanosoma cruzi activity of alkaloids isolated from Habranthus brachyandrus (Amaryllidaceae) from Argentina

BackgroundChagas disease, caused by the parasite Trypanosoma cruzi, affects over six million people worldwide, mainly in Latin American countries. Currently available drugs have variable efficacy in the chronic phase and significant side effects, so there is an urgent need for safer chemotherapeutic treatments. Natural products provide privileged structures that could serve as templates for the synthesis of new drugs. Among them, Amaryllidaceae plants have proved to be a potential natural source of therapeutical agents due to their rich diversity in alkaloids. PurposeTo identify alkaloids with anti-T. cruzi activity from Habranthus brachyandrus (Baker) Sealy (Amaryllidaceae, subfamily Amaryllidoideae) collected in Argentina. MethodsAn H. brachyandrus alkaloid extract was tested against T. cruzi, and its cytotoxicity profile was evaluated against two mammalian cell lines to ascertain its selectivity against the parasite and potential liver toxicity. It was also assessed by a stage-specific anti-amastigote assay and analysed by GC/MS to determine its alkaloid profile. The isolated alkaloids were also tested using the aforementioned assays. ResultsThe extract showed high and specific activity against T. cruzi. The alkaloids lycoramine, galanthindole, 8-O-demethylmaritidine, 8-O-demethylhomolycorine, nerinine, trisphaeridine, deoxytazettine, and tazettamide were identified by means of GC-MS. In addition, hippeastidine (also named aulicine), tazzetine, ismine, and 3-epimacronine were isolated. The alkaloid ismine was specifically active against the parasite and had low toxicity against HepG2 cells, but did not show anti-amastigote activity. ConclusionThe extract had specific anti-T. cruzi activity and the isolated alkaloid ismine was partially responsible of it. These results encourage further exploration of H. brachyandrus alkaloids in search of novel starting points for Chagas disease drug development.

Genetic and Chemical Profiling of Solenopsis spp. (Hymenoptera: Formicidae) Intercepted in Hawaii

Ants intercepted by the Hawaii Department of Agriculture in 2018 and 2019 identified as Solenopsis spp., and possibly S. invicta, were submitted to the United States Department of Agriculture-Agricultural Research Service-Center for Medical, Agricultural and Veterinary Entomology, Imported Fire Ant and Household Insects Research Unit for further taxonomic assessment using genetic and chemical methods. The 2018 sample consisted of 1 worker ant, and the 2019 sample consisted of 3 worker ants. Thoraces were used for genetic assessment and gasters were used for venom analyses. It was determined readily from genetic and chemical analyses that the intercepted ants were not S. invicta or S. richteri. Mitochondrial COX1 gene sequences were compared with previously published data from Solenopsis species. The closest match for the 2018 sample was S. xyloni, which is native to the shipment's origin (San Diego, California, USA). The 2 major alkaloids found were characteristic of the S. geminata species group. In addition, the alkaloid ratio closely matched that of S. xyloni. Therefore, the alkaloid analysis of the 2018 intercepted ant concurs with the genetic data that the ant was an S. xyloni worker. The COX1 gene sequences of the 2019 interceptions most closely matched S. geminata. The corresponding venom alkaloid profile also eliminated S. invicta and S. richteri as possibilities. However, 1 major alkaloid was unusual (Fig. 2) and normally is found only in trace amounts. We suggest the 2019 specimens are an unusual variant of S. geminata.

Identification of Veratrum Species in Pimacao Based on ITS2 Sequences and Steroidal Alkaloids by a Pseudo-Targeted Metabolomics Method.

Pimacao is a traditional Chinese folk medicine and is the main component of the famous Chinese herbal remedy “Yunnan Baiyao” for its significant analgesic activity in the treatment of wounds. Due to increases in consumption, its wild population is now difficult to find, and adulterant from the same genus has occurred. However, this is challenging to distinguish the species of Veratrum in Pimacao using dried roots and rhizomes or medicinal powder. ITS2 sequences and steroidal alkaloids by the non-targeted and pseudo-targeted metabolomics methods were taken advantage of establishing an effective identification method. Based on the ITS2 sequence, metabolite profiling of steroidal alkaloids and morphological characteristics, the classification of two distinct subspecies in V. mengzeanum has been reinforced. In addition, the new subspecies V. mengzeanum subsp. phuwae was collected in China for the first time. The ITS2 sequence could be used in the identification of V. taliense, V. mengtzeanum, V. stenophyllum, and V. nigrum, but is insufficient for intraspecific identification. Simultaneously, 147 variables were labeled by non-targeted analysis accomplished utilizing an ultra-high-performance liquid chromatography electrospray ionization orbitrap tandem mass spectrometry (UPLC-ESI-QE-Orbitrap-MS) system consisting of an Orbitrap QE HF-X. Followed by a pseudo-targeted analysis method developed for the Qtrap 6500-plus mass spectrometry system coupled with an ESI source, 29 labeled steroidal alkaloids detected by the MRM mode could distinguish between four species. Notably, 25 labeled steroidal alkaloids could distinguish between three closely related species. These have the potential to be used as markers for identification. Furthermore, there were several variables with statistical differences between two subspecies of V. mengtzeanum and populations of V. taliense, V. mengtzeanum, and V. stenophyllum.

Green Extraction-Assisted Pseudo-Targeted Profile of Alkaloids in Lotus Seed Epicarp Based on UPLC-QTOF MS with IDA.

Lotus seed epicarp, a byproduct of lotus, is commonly discarded directly or burned in the cropland, resulting in waste of resources and environmental pollution. In this work, a green ultrasonic-assisted extraction method with ethyl lactate as the extraction solvent was established to extract alkaloids from lotus seed epicarp. The extraction conditions were optimized by response surface methodology. Under the optimal extraction conditions, the extraction of alkaloids from 1 g lotus seed epicarp was accomplished with only 10 mL of extraction solvent within 15 min. Combined with ultrahigh-performance liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry with information-dependent acquisition mode, a total of 42 alkaloids were annotated in the lotus seed epicarp extracts. Among them, 39 alkaloids were reported in lotus seed epicarp for the first time. According to quantitative analysis, the distributions and trends of alkaloids in the lotus seed epicarp were found to be similar to those of lotus leaves. The five growth stages of lotus seed epicarp could be successfully distinguished based on the ten representative alkaloids. This study demonstrates that ultrasonic-assisted extraction with ethyl lactate as extractant solvent was efficient in the extraction of alkaloids from lotus seed epicarp, which is a potential renewable resource of bioactive ingredients.

Microsatellites, morphological, and alkaloids characterization of Zephyranthes fosteri and Z. alba (Amaryllidaceae): Allopatric populations

Zephyranthes (Amaryllidaceae) is a taxonomically complex genus due to the frequent overlap of interspecific morphological variation. In Mexico, Z. alba and Z. fosteri are herbaceous plants that, when distributed in sympatry, generate individuals with complex patterns of morphological variation, leading to taxonomic confusion. Therefore, it is necessary to first characterize these species in allopatric populations. In this contribution, molecular, morphological, and alkaloid profiles were used to characterize both species in allopatric sites. Our results show that Z. alba and Z. fosteri allopatric populations are two well-defined genetic and morphological groups. Flower-related characters were the ones that best allowed us to distinguish between species. In a similar fashion, the alkaloid profile showed remarkable differences among species: four alkaloids were specific to Z. alba and five to Z. fosteri. Lycorine (43.3-88.8%) and galanthamine (87.7-91.4%) were the most abundant alkaloids for each species, respectively. In conclusion, Z. fosteri and Z. alba exhibit noticeable differences when distributed in allopatry. In addition, Z. fosteri has greater genetic and phenotypic plasticity compared to Z. alba, which could be related to the former's ability to colonize new habitats. Finally, the molecular, genetic and chemical markers developed here will provide a framework to further studies aiming to explore if hybridization among Z. alba and Z. fosteri occurs in sympatric populations.

Comparative Analysis of Purine Alkaloids and Main Quality Components of the Three Camellia Species in China.

Tea (Camellia sinensis var. sinensis) is a widely consumed caffeine-containing beverage, however the Camellia genus also includes other species, which are consumed as tea in their local growing regions. Presently, HPLC analysis assessed 126 unique Camellia germplasms belonging to three Camellia species, C. sinensis var. pubilimba Chang (Csp), C. gymnogyna Chang (CgC) and C. crassicolumna Chang (CcC). Theobromine was the predominant purine alkaloid in all species, representing over 90% of purine alkaloids in Csp and CgC, and 50% in CcC. Significant variability existed in purine alkaloid patterns both between and within species, and some germplasms possessed highly unique alkaloid profiles. Sensory evaluation and quality composition analysis of green tea products produced from the three Camellia species suggested their unsuitability for use in tea production due to their unpalatable flavor. The results of this study revealed the differences in purine alkaloids and main quality components between Camellia species and tea, which contributed to understand why tea, rather than other Camellia species, has become a popular beverage in the world after long-term artificial selection. In addition, unique alkaloid profiles suggest usefulness of these germplasm resources in future breeding of decaffeinated tea plant varieties and alkaloid metabolism research.

Alkaloid Profiling, Anti-Enzymatic and Antiproliferative Activity of the Endemic Chilean Amaryllidaceae Phycella cyrtanthoides

This research aims to identify the alkaloid profile and to evaluate the enzyme inhibitory potential and antiproliferative effects of the Amaryllidaceae plant Phycella cyrtanthoides. The alkaloid extracts from bulbs and leaves were analyzed using ultrahigh performance liquid chromatography orbitrap mass spectrometry (UHPLC-Orbitrap-MS) analysis. A total of 70 alkaloids were detected in the P. cyrtanthoides’ extracts. The enzyme inhibition potential against cholinesterases (AChE: acetylcholinesterase, and BChE butyrylcholinesterase) and tyrosinase were studied. Bulbs displayed the best IC50 values against AChE (4.29 ± 0.03 µg/mL) and BChE (18.32 ± 0.03 µg/mL). These results were consistent with docking experiments with selected major compounds in the active sites of enzymes, while no activity was observed against tyrosinase enzyme. Antiproliferative effects were investigated against human cervical (HeLa), lung (A549, SW1573), colon (WiDr), and breast (HBL-100, T-47D) tumor cell lines. Bulbs and leaves were active in all cell lines (GI50 < 2.5 µg/mL). These findings suggest that the endemic Chilean plant P. cyrtanthoides contains diverse types of bioactive alkaloids with antiproliferative activities and inhibitory effects with potential therapeutic applications for neurodegenerative diseases

Insights into opium poppy (Papaver spp.) genetic diversity from genotyping-by-sequencing analysis

Opium poppy (Papaver somniferum) is one of the world’s oldest medicinal plants and a versatile model system to study secondary metabolism. However, our knowledge of its genetic diversity is limited, restricting utilization of the available germplasm for research and crop improvement. We used genotyping-by-sequencing to investigate the extent of genetic diversity and population structure in a collection of poppy germplasm consisting of 91 accessions originating in 30 countries of Europe, North Africa, America, and Asia. We identified five genetically distinct subpopulations using discriminate analysis of principal components and STRUCTURE analysis. Most accessions obtained from the same country were grouped together within subpopulations, likely a consequence of the restriction on movement of poppy germplasm. Alkaloid profiles of accessions were highly diverse, with morphine being dominant. Phylogenetic analysis identified genetic groups that were largely consistent with the subpopulations detected and that could be differentiated broadly based on traits such as number of branches and seed weight. These accessions and the associated genotypic data are valuable resources for further genetic diversity analysis, which could include definition of poppy core sets to facilitate genebank management and use of the diversity for genetic improvement of this valuable crop.