High-Resolution Liquid Chromatography-Mass Spectrometry-Based Metabolomics for the Classification of Chuquiraga (Barnadesioideae, Asteraceae): New Phenylpropanoid Derivatives as Chemical Markers for Chuquiraga spinosa.

Despite their relatively poorly investigated phytochemistry, species of the genus Chuquiraga are widely commercialized. The present study reports the use of a high-resolution liquid chromatography-mass spectrometry-based metabolomics approach coupled with exploratory and supervised multivariate statistical analyses for species classification and chemical marker identification of four species of Chuquiraga (C. jussieui, C. weberbaueri, C. spinosa, and Chuquiraga sp.) from Ecuador and Peru. Based on these analyses, a high percentage of correct classifications (87% to 100%) allowed the prediction of the taxonomic identity of Chuquiraga species. Through the metabolite selection process, several key constituents with the potential to be chemical markers were identified. Samples of C. jussieui displayed alkyl glycosides and triterpenoid glycosides as discriminating metabolites, while Chuquiraga sp. displayed high concentrations of p-hydroxyacetophenone, p-hydroxyacetophenone 4-O-glucoside, p-hydroxyacetophenone 4-O-(6-O-apiosyl)-glucoside, and quinic acid ester derivatives as the main metabolites. Caffeic acid was characteristic for C. weberbaueri samples, whereas C. spinosa displayed higher concentrations of the following new phenylpropanoid ester derivatives: 2-O-caffeoyl-4-hydroxypentanedioic acid (24), 2-O-p-coumaroyl-4-hydroxypentanedioic acid (34), 2-O-feruloyl-4-hydroxypentanedioic acid (46), 2,4-O-dicaffeoylpentanedioic acid (71), and 2-O-caffeoyl-4-O-feruloylpentanedioic acid (77).

Constituents of Croton megalocarpus with Potential Anti-HIV Activity.

Reported herein is an anti-HIV monochlorinated compound, 1ß-acetoxy-3ß-chloro-5α,6α-dihydroxycrotocascarin L (1), of the rare crotofolane diterpenoid class. Compound 1, a suspected artifact of extraction, along with the previously undescribed 11ß-acetoxycrotocascarin L (2) and a known compound, crotocascarin K (3), were isolated from the bark of Croton megalocarpus, a Kenyan oil-producing seed crop. Compounds 1 and 3 inhibited HIV-1 replication with IC50 values of 28 and 5.5 nM, respectively. Furthermore, both compounds lacked cytotoxicity toward MT-4 cells and FM-55-M1 cells at concentrations of up to 50 µM. Compounds 1 and 3 were both found to inhibit HIV-1 protease.

Fundamental Chemistry of Essential Oils and Volatile Organic Compounds, Methods of Analysis and Authentication.

The current text provides a comprehensive introduction to essential oils, their biosynthesis, naming, analysis, and chemistry. Importantly, this text quickly brings the reader up to a level of competence in the authentication of essential oils and their components. It gives detailed descriptions of enantiomers and other forms of stereoisomers relevant to the study of natural volatiles and essential oils. The text also describes GC-MS work and provides tips on rapid calculation of arithmetic indices, how to interpret suggested names from the NIST mass spectral library, and what additional efforts are required to validate essential oils and defeat sophisticated adulteration tactics. In brief, essential oils are mixtures of volatile organic compounds that were driven out of the raw plant material in distillation, condensed into an oil that is strongly aroma emitting, and collected in a vessel as the top layer (uncommonly bottom layer) of two phase separated liquids: oil and water. Essential oils commonly include components derived from two biosynthetic groups, being terpenes (monoterpenes, sesquiterpenes and their derivatives) and phenylpropanoids (aromatic ring with a propene tail). The current text provides details of how terpenes and phenylpropanoids are further categorised according to their parent skeleton, then recognised by the character of oxidation, which may be from oxygen, nitrogen, or sulphur, or the presence/absence of a double bond. The essential oil's science niche is an epicentre of individuals from diverse backgrounds, such as aromatherapy, pharmacy, synthetic and analytical chemistry, or the hobbyist. To make the science more accessible to the curious student or researcher, it was necessary to write this fundamentals-level introduction to the chemistry of essential oils (i.e., organic chemistry in the context of essential oils), which is herein presented as a comprehensive and accessible overview. Lastly, the current review constitutes the only resource that highlights common errors and explains in simplistic detail how to correctly interpret GC-MS data then accurately present the respective chemical information to the wider scientific audience. Therefore, detailed study of the contents herein will equip the individual with prerequisite knowledge necessary to effectively analyse an essential oil and make qualified judgement on its authenticity.

Chemical Diversity of Flavan-3-Ols in Grape Seeds: Modulating Factors and Quality Requirements.

Grape seeds are a rich source of flavan-3-ol monomers, oligomers, and polymers. The diverse profile of compounds includes mainly B-type procyanidins (especially C4→C8 linked molecules) and the key monomers, catechin, and epicatechin that are positively implicated in the 'French Paradox'. Today grape seed nutraceuticals have become a multi-million-dollar industry. This has created incentives to elucidate the variations in chemistry across cultivars, to identify signs of adulteration, and to understand the intrinsic and extrinsic factors controlling the expression of metabolites in the seeds' metabolome. This review provides a critical overview of the existing literature on grape seed chemistry. Although the biosynthetic pathways for polymeric procyanidins in seeds have not yet been explained, abiotic factors have been shown to modulate associated genes. Research of extrinsic factors has demonstrated that the control of procyanidin expression is strongly influenced, in order of importance, by genotype (species first, then variety) and environment, as claimed anecdotally. Unfortunately, research outcomes on the effects of abiotic factors have low certainty, because effects can be specific to genotype or variety, and there is limited control over physical metrics in the field. Thus, to gain a fuller understanding of the effects of abiotic factors and biosynthetic pathways, and realise potential for optimisation, a more fundamental research approach is needed. Nevertheless, the current synthesis offers insight into the selection of species or varieties according to the profile of polyphenols, as well as for optimisation of horticultural practices, with a view to produce products that contain the compounds that support health claims.

HIV-1 Integrase Inhibitory Effects of Major Compounds Present in CareVid™: An Anti-HIV Multi-Herbal Remedy.

In our continued study on the anti-HIV activity of compounds present in CareVidTM, we report the HIV-1 integrase ((HIV-1 IN) inhibitory effects of pellitorine (1), oleuropein (2), magnoflorine (3), crotepoxide (4), ent-kaurane-16ß,17-diol (5), crotocorylifuran (6), lupeol (7), betulin (8), and ellagic acid (9) in an in vitro enzyme assay, and in an in silico study. Ellagic acid, pellitorine, lupeol, and betulin showed an in vitro percentage inhibition against HIV-1 IN of 21.1%, 19.0%, 18.5%, and 16.8%, respectively, at a standard concentration of 25 µg/mL. However, from a pharmacokinetic perspective, ellagic acid has poor bioavailability, due to rapid elimination in metabolism in the gut microbiome. It was postulated that known gut catabolites of ellagic acid, urolithin A (10) and urolithin B (11) could be more promising candidates in exploring the anti-HIV activity of ellagic acid-rich medicinal species consumed orally. On the contrary, urolithin A and urolithin B demonstrated lower activity with comparison to ellagic acid. The binding affinity of compounds 1-9, urolithin A, and urolithin B against the catalytic domain of HIV-1 IN was also explored by in silico methods. Docking studies showed oleuropein as the best candidate, with a predicted energy of binding of ΔG -5.81 kcal/mol, while ellagic acid showed moderate predicted inhibition (ΔG -4.38 kcal/mol) caused by the interaction between the carbonyl and the key Mg2+ ion in the active site.

Emergence of methicillin resistance predates the clinical use of antibiotics.

The discovery of antibiotics more than 80 years ago has led to considerable improvements in human and animal health. Although antibiotic resistance in environmental bacteria is ancient, resistance in human pathogens is thought to be a modern phenomenon that is driven by the clinical use of antibiotics1. Here we show that particular lineages of methicillin-resistant Staphylococcus aureus-a notorious human pathogen-appeared in European hedgehogs in the pre-antibiotic era. Subsequently, these lineages spread within the local hedgehog populations and between hedgehogs and secondary hosts, including livestock and humans. We also demonstrate that the hedgehog dermatophyte Trichophyton erinacei produces two ß-lactam antibiotics that provide a natural selective environment in which methicillin-resistant S. aureus isolates have an advantage over susceptible isolates. Together, these results suggest that methicillin resistance emerged in the pre-antibiotic era as a co-evolutionary adaptation of S. aureus to the colonization of dermatophyte-infected hedgehogs. The evolution of clinically relevant antibiotic-resistance genes in wild animals and the connectivity of natural, agricultural and human ecosystems demonstrate that the use of a One Health approach is critical for our understanding and management of antibiotic resistance, which is one of the biggest threats to global health, food security and development.

HIV-1 Reverse Transcriptase Inhibition by Major Compounds in a Kenyan Multi-Herbal Composition (CareVid™): In Vitro and In Silico Contrast.

CareVid is a multi-herbal product used in southwest Kenya as an immune booster and health tonic and has been anecdotally described as improving the condition of HIV-positive patients. The product is made up of roots, barks and whole plant of 14 African medicinal plants: Acacia nilotica (L.) Willd. ex Delile (currently, Vachelia nilotica (L.) P.J.H Hurter & Mabb.), Adenia gummifera (Harv.) Harms, Anthocleista grandiflora Gilg, Asparagus africanus Lam., Bersama abyssinica Fresen., Clematis hirsuta Guill. & Perr., Croton macrostachyus Hochst. ex Delile, Clutia robusta Pax (accepted as Clutia kilimandscharica Engl.), Dovyalis abyssinica (A. Rich.) Warb, Ekebergia capensis Sparm., Periploca linearifolia Quart.-Dill. & A. Rich., Plantago palmata Hook.f., Prunus africana Hook.f. Kalkman and Rhamnus prinoides L'Her. The objective of this study was to determine the major chemical constituents of CareVid solvent extracts and screen them for in vitro and in silico activity against the HIV-1 reverse transcriptase enzyme. To achieve this, CareVid was separately extracted using CH2Cl2, MeOH, 80% EtOH in H2O, cold H2O, hot H2O and acidified H2O (pH 1.5-3.5). The extracts were analysed using HPLC-MS equipped with UV diode array detection. HIV-1 reverse transcriptase inhibition was performed in vitro and compared to in silico HIV-1 reverse transcriptase inhibition, with the latter carried out using MOE software, placing the docking on the hydrophobic pocket in the subdomain of p66, the NNRTI pocket. The MeOH and 80% EtOH extracts showed strong in vitro HIV-1 reverse transcriptase inhibition, with an EC50 of 7 µg·mL-1. The major components were identified as sucrose, citric acid, ellagic acid, catechin 3-hexoside, epicatechin 3-hexoside, procyanidin B, hesperetin O-rutinoside, pellitorine, mangiferin, isomangiferin, 4-O-coumaroulquinic acid, ellagic acid, ellagic acid O-pentoside, crotepoxide, oleuropein, magnoflorine, tremulacin and an isomer of dammarane tetrol. Ellagic acid and procyanidin B inhibited the HIV-1 reverse transcription process at 15 and 3.2 µg/mL-1, respectively. Docking studies did not agree with in vitro results because the best scoring ligand was crotepoxide (ΔG = -8.55 kcal/mol), followed by magnoflorine (ΔG = -8.39 kcal/mol). This study showed that CareVid has contrasting in vitro and in silico activity against HIV-1 reverse transcriptase. However, the strongest in vitro inhibitors were ellagic acid and procyanidin B.

Barbeya oleoides Leaves Extracts: In Vitro Carbohydrate Digestive Enzymes Inhibition and Phytochemical Characterization.

This study investigated the in vitro inhibitory potential of different solvent extracts of leaves of Barbeya oleoides on key enzymes related to type 2 diabetes mellitus (α-glucosidase and α-amylase) in combination with an aggregation assay (using 0.01% Triton X-100 detergent) to assess the specificity of action. The methanol extract was the most active in inhibiting α-glucosidase and α-amylase, with IC50 values of 6.67 ± 0.30 and 25.62 ± 4.12 µg/mL, respectively. However, these activities were significantly attenuated in the presence of 0.01% Triton X-100. The chemical analysis of the methanol extract was conducted utilizing a dereplication approach combing LC-ESI-MS/MS and database searching. The chemical analysis detected 27 major peaks in the negative ion mode, and 24 phenolic compounds, predominantly tannins and flavonol glycosides derivatives, were tentatively identified. Our data indicate that the enzyme inhibitory activity was probably due to aggregation-based inhibition, perhaps linked to polyphenols.

Cyanogenic Derivatives as Chemical Markers for the Authentication of Commercial Products of Bamboo Shoots.

The authentication of bamboo shoots found in the marketplace is complex because the chemical profile of processed and unprocessed material is different. During processing, heat derivatives of the potentially toxic cyanogenic glycoside taxiphyllin are produced. Here, we report the isolation and structure elucidation of the two major diarylbutenedinitrile derivatives, which are cis and trans isomers of the rare 2,3-bis(4-hydroxyphenyl)but-2-enedinitrile from a commercial extract of bamboo shoots. These compounds, absent in fresh bamboo shoots, were produced by boiling the shoots of Bambusa vulgaris and were associated with a decrease in levels of taxiphyllin. Furthermore, (E)-2,3-bis(4-hydroxyphenyl)but-2-enedinitrile was quantified in all 16 of the commercial products tested. Its abundance was found to be highly variable, ranging from 1 to 3 mg/g in preserved bamboo shoots and from 10 to 160 mg/mL in commercial aqueous extracts. Of the 15 authenticated bamboo samples tested for taxiphyllin, it was found only in the shoots of B. vulgaris and Gigantochloa verticillata, which represent two edible bamboo species. Our results indicate that diarylbutenedinitriles can be used as markers for the authentication of boil-processed bamboo shoots obtained from taxiphyllin-containing edible species and organs.

Effect of the Andean Geography and Climate on the Specialized Metabolism of Its Vegetation: The Subtribe Espeletiinae (Asteraceae) as a Case Example.

The Andean mountains are 'center stage' to some of the most spectacular examples of plant diversifications, where geographic isolation and past climatic fluctuations have played a major role. However, the influence of Andean geography and climate as drivers of metabolic variation in Andean plants is poorly elucidated. Here, we studied the influence of those factors on the metabolome of the subtribe Espeletiinae (Asteraceae) using liquid chromatography coupled to high-resolution mass spectrometry data of over two hundred samples from multiple locations. Our results demonstrate that metabolic profiles can discriminate Espeletiinae taxa at different geographic scales, revealing inter- and intraspecific metabolic variations: at the country level, segregation between Colombian and Venezuelan taxa was observed; regionally, between páramo massifs; and locally, between páramo complexes. Metabolic differences in Espeletiinae were mainly explained by geographic isolation, although differences in taxonomic genera, temperature, and elevation, were also important. Furthermore, we found that different species inhabiting the same páramo complex showed stronger chemical similarities than the same species at different locations, corroborating that geographic isolation represents the main driver of metabolic change in Espeletiinae. The current study serves as a starting point to fill in the gaps in how Andean geography and climate have shaped the metabolism of its vegetation and reveal the potential of untargeted metabolomics to study the environmental physiology of plants.

The Diversity of Volatile Compounds in Australia's Semi-Desert Genus Eremophila (Scrophulariaceae).

Australia's endemic desert shrubs are commonly aromatic, with chemically diverse terpenes and phenylpropanoids in their headspace profiles. Species from the genus Eremophila (Scrophulariaceae ex. Myoporaceae) are the most common, with 215 recognised taxa and many more that have not yet been described, widely spread across the arid parts of the Australian continent. Over the years, our research team has collected multiple specimens as part of a survey to investigate the chemical diversity of the genus and create leads for further scientific enquiry. In the current study, the diversity of volatile compounds is studied using hydrodistilled essential oils and leaf solvent extracts from 30 taxa. Several rare terpenes and iridoids were detected in chemical profiles widely across the genus, and three previously undescribed sesquiterpenes were isolated and are assigned by 2D NMR-E-11(12)-dehydroisodendrolasin, Z-11-hydroxyisodendrolasin and 10-hydroxydihydro-α-humulene acetate. Multiple sampling from Eremophila longifolia, Eremophila arbuscular, Eremophila latrobei, Eremophila deserti, Eremophila sturtii, Eremophila oppositifolia and Eremophila alternifolia coneys that species in Eremophila are highly chemovariable. However, taxa are generally grouped according to the expression of (1) furanosesquiterpenes, (2) iridoids or oxides, (3) mixtures of 1 and 2, (4) phenylpropanoids, (5) non-furanoid terpenes, (6) mixtures of 4 and 5, and less commonly (7) mixtures of 1 and 5. Furthermore, GC-MS analysis of solvent-extracted leaves taken from cultivated specimens conveys that many heavier 'volatiles' with lower vapour pressure are not detected in hydrodistilled essential oils and have therefore been neglected in past chemical studies. Hence, our data reiterate that chemical studies of the genus Eremophila will continue to describe new metabolites and that taxon determination has limited predictive value for the chemical composition.

Prostanthera (Lamiaceae) as a 'Cradle of Incense': Chemophenetics of Rare Essential Oils from Both New and Forgotten Australian 'Mint Bush' Species.

The highly aromatic Australian mint bushes from the genus Prostanthera Labill. produce a high yield of essential oil on hydrodistillation. Together with its rich history, horticultural potential, iconic flowers, and aromatic leaves, it achieves high ornamental and culinary value. Species in the genus express highly diverse and chemically unique essential oils that demonstrate intra- and inter-specific patterns that have inspired taxonomic reinterpretation for over a hundred years. Previous studies have conveyed that phenoplastic expression of volatiles creates chemotypes within taxa, adding complexity to chemophenetic exploration. The current study chemically characterised essential oils from 64 highly aromatic specimens, representative of 25 taxa, giving yields as high as >2% g/g. The chemical profiles of essential oils are diverse, but generally include 1,8-cineole and signatory compounds such as sesquiterpene oxides, caryophyllene oxide, kessane and cis-dihydroagarofuran; sesquiterpene alcohols, globulol, epiglobulol, maaliol, prostantherol, spathulenol and ledol; and monoterpene derivatives of common scaffolds, borneol, bornyl acetate, bornanone, linalool and linalyl acetate. As in previous studies, analysis of chemical data confirms that the chemistry strongly agrees with taxonomic classifications. Importantly, as in classical taxonomy, the current chemical study complemented morphological analysis but conveys chemovariation, obscuring the taxonomic agreement. Nevertheless, variation within taxa may be due to environmental factors, meaning that cultivation of species in gardens will create different chemical profiles as compared to those published here.

Feature-Based Molecular Networking to Target the Isolation of New Caffeic Acid Esters from Yacon (Smallanthus sonchifolius, Asteraceae).

Smallanthus sonchifolius (yacon) is an edible tuberous Andean shrub that has been included in the diet of indigenous people since before recorded history. The nutraceutical and medicinal properties of yacon are widely recognized, especially for the improvement of hyperglycemic disorders. However, the chemical diversity of the main bioactive series of caffeic acid esters has not been explored in detail. In this metabolomics study, we applied the latest tools to facilitate the targeted isolation of new caffeic acid esters. Using liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS), we analyzed extracts from different organs (roots, vascular tissues of the stems, stem epidermis, leaves, bracts, and ray flowers) and followed a feature-based molecular networking approach to characterize the structural diversity of caffeic acid esters and recognize new compounds. The analysis identified three potentially new metabolites, one of them confirmed by isolation and full spectroscopic/spectrometric assignment using nuclear magnetic resonance (NMR), high-resolution mass spectrometry (HRMS), and MS/MS. This metabolite (5-O-caffeoyl-2,7-anhydro-d-glycero-ß-d-galacto-oct-2-ulopyranosonic acid), along with eight known caffeic acid esters, was isolated from the roots and stems. Furthermore, based on detailed tandem MS analyses, we suggest that the two isomeric monocaffeoyl-2,7-anhydro-2-octulopyranosonic acids found in yacon can be reliably distinguished based on their characteristic MS2 and MS3 spectra. The outcome of the current study confirms the utility of feature-based molecular networking as a tool for targeted isolation of previously undescribed metabolites and reveals the full diversity of potentially bioactive metabolites from S. sonchifolius.

Metabolomic and Gene Expression Studies Reveal the Diversity, Distribution and Spatial Regulation of the Specialized Metabolism of Yacón (Smallanthus sonchifolius, Asteraceae).

Smallanthus sonchifolius, also known as yacón, is an Andean crop species commercialized for its nutraceutical and medicinal properties. The tuberous roots of yacón accumulate a diverse array of probiotic and bioactive metabolites including fructooligosaccharides and caffeic acid esters. However, the metabolic diversity of yacón remains unexplored, including the site of biosynthesis and accumulation of key metabolite classes. We report herein a multidisciplinary approach involving metabolomics, gene expression and scanning electron microscopy, to provide a comprehensive analysis of the diversity, distribution and spatial regulation of the specialized metabolism in yacón. Our results demonstrate that different metabolic fingerprints and gene expression patterns characterize specific tissues, organs and cultivars of yacón. Manual inspection of mass spectrometry data and molecular networking allowed the tentative identification of 71 metabolites, including undescribed structural analogues of known bioactive compounds. Imaging by scanning electron microscopy revealed the presence of a new type of glandular trichome in yacón bracts, with a distinctive metabolite profile. Furthermore, the high concentration of sesquiterpene lactones in capitate glandular trichomes and the restricted presence of certain flavonoids and caffeic acid esters in underground organs and internal tissues suggests that these metabolites could be involved in protective and ecological functions. This study demonstrates that individual organs and tissues make specific contributions to the highly diverse and specialized metabolome of yacón, which is proving to be a reservoir of previously undescribed molecules of potential significance in human health.

Metabolomic and gene expression approaches reveal the developmental and environmental regulation of the secondary metabolism of yacón (Smallanthus sonchifolius, Asteraceae).

Acting as chemical defense or signaling compounds, secondary metabolites (SMs) play an essential role in the evolutionary success of many angiosperm plant families. However, the adaptive advantages that SMs confer, and the influence of environmental and developmental factors on SMs expression, remains poorly understood. A study of taxa endemic to the variable Andean climate, using a metabolomics approach, may provide further insight. By analyzing gene expression patterns and metabolic fingerprints, we report herein the developmental and environmental regulation of the secondary metabolism of Smallanthus sonchifolius (yacón), a medicinal Andean plant. Our results demonstrate a clear developmental stage dependent regulation of the secondary metabolism of yacón leaves wherein the metabolic diversity increases with plant age. However, environmental factors seem to regulate biosynthetic pathways, creating differences in the expression of chemical classes, pointing to an association between transcription levels of relevant genes and the relative amounts of more than 40 different metabolites. This study suggests that the secondary metabolism of yacón is regulated by a complex interplay between environmental factors and developmental stage and provides insight into the regulatory factors and adaptive roles of SMs in Andean taxa.

Genetic and chemical diversity among yacon [Smallanthus sonchifolius (Poepp. et Endl.) H. Robinson] accessions based on iPBS markers and metabolomic fingerprinting.

The present study is focused on the characterization of yacon [Smallanthus sonchifolius (Poepp. et Endl.) H. Robinson] accessions from different geographic origins (Bolivia, Ecuador, and Peru) by iPBS markers and metabolomic fingerprinting. The results showed that the number of amplified polymorphic fragment levels ranged from 20 up to 27 with a level of polymorphism ranging from 80 to 100%. Five of the iPBS primers used in this study provided no specific banding pattern able to discriminate between the different yacon accessions. However, two iPBS primer pairs were able to separate Peru accessions from those of Ecuador and Bolivia. The UPLC-HRMS/MS-based metabolomic fingerprinting showed highly similar metabolomic fingerprints characterized by the accumulation of high quantities of sesquiterpene lactones and diterpenes, but no apparent geographic clustering. The present study demonstrates that yacon accessions from different geographical origins maintained ex situ (in the Czech Republic) present a rather low chemical and genetic diversity.

Biogeography shaped the metabolome of the genus Espeletia: a phytochemical perspective on an Andean adaptive radiation.

The páramo ecosystem has the highest rate of diversification across plant lineages on earth, of which the genus Espeletia (Asteraceae) is a prime example. The current distribution and molecular phylogeny of Espeletia suggest the influence of Andean geography and past climatic fluctuations on the diversification of this genus. However, molecular markers have failed to reveal subtle biogeographical trends in Espeletia diversification, and metabolomic evidence for allopatric segregation in plants has never been reported. Here, we present for the first time a metabolomics approach based on liquid chromatography-mass spectrometry for revealing subtle biogeographical trends in Espeletia diversification. We demonstrate that Espeletia lineages can be distinguished by means of different metabolic fingerprints correlated to the country of origin on a global scale and to the páramo massif on a regional scale. Distinctive patterns in the accumulation of secondary metabolites according to the main diversification centers of Espeletia are also identified and a comprehensive phytochemical characterization is reported. These findings demonstrate that a variation in the metabolic fingerprints of Espeletia lineages followed the biogeography of this genus, suggesting that our untargeted metabolomics approach can be potentially used as a model to understand the biogeographic history of additional plant groups in the páramo ecosystem.

Chemical characterization of two morphologically related Espeletia (Asteraceae) species and chemometric analysis based on essential oil components

ABSTRACT In this study, a comprehensive phytochemical characterization of two morphologically related species from the genus Espeletia Mutis ex Bonpl., namely, Espeletia grandiflora Humb. & Bonpl. and Espeletia killipii Cuatrec., Asteraceae, has been performed by gas chromatography coupled to flame ionization detection, gas chromatography coupled to mass spectrometry and ultra-high performance liquid chromatography coupled to ultraviolet and high-resolution mass spectrometry. Analysis of ethanol extracts (70%, v/v) from leaves and concomitant compound dereplication allowed the identification of major peaks, most of them new reports for the genus Espeletia or the subtribe Espeletiinae. Chemical characterization of resins essential oils indicated several similarities and differences between both species and from other members of the subtribe. Chemometric analysis (hierarchical clustering analysis and orthogonal partial least-squares discriminant analysis) applied to the essential oil composition of 31 species from Espeletiinae furthermore allowed the identification of three primary clusters correlated with the taxonomy. Hence, this study underscored qualitative and semiquantitative differences between the chemical composition of leaves and resins of E. grandiflora and E. killipii, provided information on chemotaxonomy and described the presence of different trends in the essential oil composition from species of Espeletiinae.

Actividad antibacteriana de extractos y fracciones de hojas de Siparuna sessiliflora Kunth A. DC. (limoncillo) / Anti-bacterial action of extracts and fractions from Siparuna sessiliflora Kunth A. DC (limoncillo)

Introducción: Siparuna sessiliflora Kunth A. DC. es una especie vegetal comúnmente conocida en Colombia como limoncillo, utilizada por parte de varias comunidades indígenas para tratar diferentes problemas de salud. A pesar de contar con diversos registros de usos etnobotánicos, esta especie ha sido poco estudiada desde el punto de vista fitoquímico o de su actividad biológica. Objetivos: evaluar la actividad antibacteriana de los extractos y las fracciones obtenidos a partir de las hojas de Siparuna sessiliflora sobre Bacillus subtilis, Staphylococcus aureus, Escherichia coli y Pseudomonas aeruginosa. Métodos: el material vegetal fue extraído por maceración en frío con éter de petróleo y etanol 96 por ciento; estos extractos se fraccionaron por métodos cromatográficos y partición líquido/líquido; su actividad antibacteriana se evaluó por el método de difusión en gel por perforación en placa. Posteriormente, se identificaron los principales compuestos responsables de la bioactividad por medio de cromatografía de gases acolada a espectrometría de masas. Resultados: se encontró que las fracciones de alcaloides Alk 1 y Alk 2 presentaron actividad frente a B. subtilis, S. aureus y E. coli. Adicionalmente, la subfracción Alk 2A fue la más activa frente a B. subtilis. Se identificaron sobre todo alcaloides isoquinolínicos y un derivado del ácido cinámico como posibles compuestos responsables de la bioactividad. Conclusiones: se reportó la actividad antibacteriana de S. sessiliflora frente a bacterias grampositivas y gramnegativas, y se generó nuevo conocimiento sobre su fitoquímica que corrobora su uso tradicional en la cura de cuadros infecciosos.

Introduction: Siparuna sessiliflora Kunth A. DC. is a plant species popularly known in Colombia as «limoncillo¼ and used by several indigenous communities for treating different health disorders. Despite the many records of its ethnobotanical uses, this species has been poorly studied from a phytochemical or biological standpoint. Objectives: to evaluate the antibacterial activity of the extracts and fractions obtained from Siparuna sessiliflora´s leaves against Bacillus subtilis, Staphylococcus aureus, Escherichia coli and Pseudomonas aeruginosa. Methods: the plant material was extracted by cold maceration with petroleum ether and 96 percent ethanol. Those extracts were fractionated by chromatographic methods and liquid-liquid partitioning; additionally, its antibacterial activity was assessed by gel diffusion. Subsequently, there were identified the main bioactive compounds by gas chromatography-mass spectrometry. Results: it was found that the alkaloidal fractions Alk 1 and Alk 2 were active against B. subtilis, S. aureus and E. coli. Additionally, the subfraction Alk 2A was the most active against B. subtilis. Isoquinoline alkaloids and a cinnamic acid derivative were identified as possible compounds responsible for bioactivity. Conclusions: the antibacterial activity of S. sessiliflora against Gram positive and Gram negative bacteria was reported, so the new knowledge about its phytochemistry endorses its traditional use in the treatment of infections.