The agavins (Agave carbohydrates) story.

Fructans, are carbohydrates defined as fructose-based polymers with countable degree of polymerization (DP) ranging so far from DP3 to DP60. There are different types of fructans depending on their molecular arrangement. They are categorized as linear inulins and levans, neoseries of inulin and levan, branched graminans, and highly branched neofructans, so called agavins (Agave carbohydrates). It is worth to note that agavins are the most recently described type of fructans and they are also the most complex ones. The complexity of these carbohydrates is correlated to their various isomers and degree of polymerization range, which is correlated to their multifunctional application in industry and human health. Here, we narrate the story of the agavins' discovery. This included their chemical characterization, their benefits, biotechnological applications, and drawbacks over human health. Finally, a perspective of the study of agavins and their interactions with other metabolites through metabolomics is proposed.

Agave amica a potential model for the study of agavins metabolism.

Fructans found in agave are called agavins, highly branched neo-fructans. They are essential on the yield and quality of Tequila production. The need for agave specimens with higher accumulation of agavins became essential before the growing demand of such products. To get such specimens, understanding agavins metabolism is a quintessential requirement. For this, a more efficient biological model is required. The recently reclassified Agave amica possesses the potential to gather the requirements for becoming such a model. Therefore, this study dealt with the characterization of carbohydrates in the bulbs of A. amica focusing on fructans. Moreover, it tested and described its feasibility as model for the accelerated study of agavins. Infrared analysis unveiled potential content of fructans in the bulbs of A. amica. Furthermore, high performance thin layer chromatography detected fructooligosaccharides. High performance anion exchange chromatography confirmed a polydisperse mixture of branched fructans. Gas chromatography-mass spectrometry analysis demonstrated agavins like structures in the bulbs of A. amica. Moreover, total fructan content and multivariate data analysis through bulb's age demonstrated their correlation. Thus, the presence of agavins, their correlation with phenology, and their technical advantages highlighted the feasibility of this species as a potential new biological model for the study of agavins' metabolism.

HPTLC-based fingerprinting: An alternative approach for fructooligosaccharides metabolism profiling.

Fructans are categorized as fructose-based metabolites with no more than one glucose in their structure. Agave species possess a mixture of linear and ramified fructans with different degrees of polymerization. Among them, fructooligosaccharides are fructans with low degree of polymerization which might be approachable by high performance thin layer chromatography (HPTLC). Thus, this study used two emblematic Agave species collected at different ages as models to explore the feasibility of HPTLC-based fingerprinting to characterize fructooligosaccharides (FOS) production, accumulation, and behavior through time. To do so, high performance anion exchange was also used as analytical reference to determine the goodness and robustness of HPTLC data. The multivariate data analysis showed separation of samples dictated by species and age effects detected by both techniques. Moreover, linear correlations between the increase of the age in agave and their carbohydrate fraction was established in both species by both techniques. Oligosaccharides found to be correlated to species and age factors, these suggest changes in specific carbohydrate metabolism enzymes. Thus, HPTLC was proven as a complementary or stand-alone fingerprinting platform for fructooligosaccharides characterization in biological mixtures. However, the type of derivatizing reagent and the extraction color channel determined the goodness of the model used to scrutinize agavin fructooligosaccharides (aFOS).

Metabolomic Investigation of Citrus latifolia and the Putative Role of Coumarins in Resistance to Black Spot Disease.

Citrus black spot (CBS) is a disease caused by the fungus Phyllosticta citricarpa that affects citrus plants, causing fruit blemish and premature drop that result in severe economic losses in commercial citrus orchards. However, CBS symptoms and effects may vary depending on the citrus species: Citrus limon (lemon) is susceptible and highly affected by the disease, while no CBS-related damage has ever been observed for Citrus latifolia (Tahiti lime), implying that it must be resistant to the disease. The difference in the response to this disease provided the opportunity to gain insight into the metabolites responsible for the resistance by comparison of the metabolomic profiles of these two citrus species. Metabolic variations of C. limon and C. latifolia inoculated with P. citricarpa were analyzed using various metabolomic-based platforms including 1H NMR for overall metabolic profiling, and LC-MS and HPTLC for targeted analysis. The 1H NMR spectra of the samples demonstrated that certain phenolics were strongly induced after pathogenic inoculation, especially in the resistant species. The induced phenolics were identified from C. latifolia by further 1H NMR, LCMS and HPTLC analysis yielding six prenylated and methoxy coumarins, i.e., 5,7-dimethoxycoumarin, 5-geranyloxy-7-methoxycoumarin, 7-geranyloxycoumarin, 8-methoxypsoralen, 5,8-dimethoxypsoralen and 5-geranyloxypsoralen. These isolated coumarins and a coumarin-rich fraction were tested against the fungal pathogen, P. citricarpa, to evaluate their activity. None of the individual coumarins exhibited a significant inhibition, while the coumarin fraction exhibited a strong antifungal activity suggesting a synergistic interaction of its components. To obtain further insight into the roles of these compounds in the plant defense, the possible mechanisms of the individual coumarins were tested using an in-silico model, the PASS Online Tool; the analysis showed that each coumarin appeared to have a unique defense mechanism, even with very slight variations in the chemical structures. The results could provide evidence of the existence of a complex plant defense mechanism consisting in a multitude of synergistic interactions between compounds.

A Theme Issue to Celebrate Professor Robert Verpoorte's 75th Birthday: "The Past, Current, and Future of Natural Products".

Dear Colleagues, [...].

Chemical Differentiation of Plant Latexes and Their Anti-herbivory Activity against Thrips Frankliniella occidentalis.

Despite the extensive studies on latex, some fundamental questions on their chemical specialization and the factors influencing this specialization have yet to be investigated. To address this issue, latexes and their bearing tissues from diverse species were profiled by 1HNMR and GC-MS. Additionally, the antiherbivory activity of these materials was tested against thrips (Frankliniella occidentalis Pergande, 1895). The multivariate data analysis showed a clear separation between latexes and leaves from the same species. Conversely, the chemical profiles of latexes from different species were highly similar, that is, they displayed much less metabolic species-specificity. These shared chemical profiles of latexes were reflected in their overall higher mortality index (80.4% ± 7.5) against thrips compared with their bearing tissues (55.5% ± 14.9). The metabolites correlated to the antiherbivory activity of latexes were triterpenoids and steroids. However, the activity could not be attributed to any single terpenoid. This discrepancy and the reduction of the latex activity after fractionation suggested a complementary effect of the compounds when in a mixture as represented by the latex. Additionally, aqueous fractions of several latexes were found to possess simple spectra, even with only 1 metabolite. These metabolites were determined to be organic acids that might be involved in the modulation of the rate of latex coagulation, potentially increasing the sealing and trapping effects of the latex.

Latex Metabolome of Euphorbia Species: Geographical and Inter-Species Variation and its Proposed Role in Plant Defense against Herbivores and Pathogens.

Based on the hypothesis that the variation of the metabolomes of latex is a response to selective pressure and should thus be affected differently from other organs, their variation could provide an insight into the defensive chemical selection of plants. Metabolic profiling was used to compare tissues of three Euphorbia species collected in diverse regions. The metabolic variation of latexes was much more limited than that of other organs. In all the species, the levels of polyisoprenes and terpenes were found to be much higher in latexes than in leaves and roots of the corresponding plants. Polyisoprenes were observed to physically delay the contact of pathogens with plant tissues and their growth. A secondary barrier composed of terpenes in latex and in particular, 24-methylenecycloartanol, exhibited antifungal activity. These results added to the well-known role of enzymes also present in latexes, show that these are part of a cooperative defense system comprising biochemical and physical elements.

Metabolic variation in Cistus monspeliensis L. ecotypes correlated to their plant-fungal interactions.

The effect of environmental factors on the chemical composition of plants eventually resulting in plant growth regulation is an age-old issue in plant biology. Nowadays, the acceleration in changes in environmental conditions (e.g. global warming) can act as an incentive to investigate their correlation with metabolic changes. In this study, Cistus monspeliensis plants grown on the island of Sardinia (Italy) were used to explore the geographical-mediated metabolic variation and its repercussion on plant-fungus interactions. Samples of different ecotypes of C. monspeliensis were collected and chemically profiled by 1H NMR and HPTLC-based metabolomics and the relationship between the variations of biological activity was examined by multivariate data analysis. The ecotypes, collected from different geographical zones and altitudes, exhibited clearly distinguishable chemical profiles, particularly in their terpene and phenolic contents. In particular, multivariate data analysis revealed several diterpenes of the labdane and clerodane series among the terpenes and methoxyflavonoids to be responsible for the differentiation. The antifungal activity of the plants was used to explore the correlation between chemical variation and biological activity. Results showed that there was a strong correlation between the metabolic profiles and the antifungal activity, revealing terpenes and methoxylated flavonoids as the main involved metabolites. This demonstrated that environmental factors can influence the chemical variation of plant ecotypes, resulting in the generation of chemotypes that are potentially adapted to their niche conditions including the plant-fungal interactions.

Proximate mechanisms of drought resistance in Phytoseiulus persimilis eggs.

Under drought stress, Phytoseiulus persimilis females are able to lay drought-resistant eggs through an adaptive maternal effect. The mechanisms making these eggs drought resistant still remain to be investigated. For this purpose, we studied the physiological differences between drought-resistant and drought-sensitive eggs. We compared the volume and the surface-area-to-volume ratio (SA:V) of the eggs, their sex ratio, their chemical composition (by gas chromatography-mass spectrometry), their internal and external structure [by scanning electron microscope (SEM) and transmission electron microscope (TEM) images], and their developmental time. Our results show that drought-resistant and drought-sensitive eggs have a different chemical composition: drought-resistant eggs contain more compatible solutes (free amino acids and sugar alcohols) and saturated hydrocarbons than drought-sensitive eggs. This difference may contribute to reducing water loss in drought-resistant eggs. Moreover, drought-resistant eggs are on average 8.4% larger in volume, and have a 2.4% smaller SA:V than drought-sensitive eggs. This larger volume and smaller SA:V, probably the result of a higher water content, may make drought-resistant eggs less vulnerable to water loss. We did not find any difference in sex ratio, internal or external structure nor developmental time between drought-resistant and drought-sensitive eggs. These results mark the first step in the understanding of the strategies and the energetic costs involved in the production of drought-resistant eggs in P. persimilis females.

Plant Latex, from Ecological Interests to Bioactive Chemical Resources.

Historically, latex-bearing plants have been regarded as important medicinal resources in many countries due to their characteristic latex ingredients. They have also often been endowed with a social or cultural significance in religious or cult rituals or for hunting. Initial chemical studies focused on the protein or peptide content but recently the interest extended to smaller molecules. Latex has been found to contain a broad range of specialized metabolites such as terpenoids, cardenolides, alkaloids, and phenolics, which are partly responsible for their antibacterial, antifungal, anthelmintic, cytotoxic, and insect-repellent activities. The diversity in biology and chemistry of latexes is supposedly associated to their ecological roles in interactions with exogenous factors. Latexes contain unique compounds that are different to those found in their bearing plants. Exploring the feasibility of plant latex as a new type of bioactive chemical resource, this review paper covers the chemical characterization of plant latexes, extending this to various other plant exudates. Also, the factors influencing this chemical differentiation and the production, transportation, and chemistry of the latex exudates are described, based on ecological and biochemical mechanisms. We also proposed a latex coagulation model involving 4 general conserved steps. Therefore, the inherent defensive origin of latexes is recognized as their most valuable character and encourages one to pay attention to these materials as alternative sources to discover metabolites with insecticidal or antimicrobial activity.

Metabolic discrimination of pine resins using multiple analytical platforms.

Resins are one of the first sites of interaction between plants and biotic and abiotic factors. Despite their evident morphological and chemical differentiation from other plant organs, the detailed correlation between resins and biological or environmental factors is not yet clear. In this study, 1H nuclear magnetic resonance (NMR), gas chromatography coupled with mass spectrometry (GC-MS) and high-performance thin-layer chromatography (HPTLC)-based profiling techniques were applied to the metabolic characterisation of plant resins of different species and season of collection, using samples from five different species that were collected during early and late spring. The 1H NMR analysis confirmed the main metabolic groups in the resins to be terpenoids and further GC-MS analysis revealed a notable chemical variation between the species and collection periods. Abies grandis displayed a significant differentiation from the other species, showing a higher number of monoterpenes. The HPTLC-based profiling method hyphenated with multivariate data analysis (MVDA) also showed a clear separation confirming the GC-MS terpenoidal profiling results. Additionally, the unknown compounds were obtained by preparative TLC for identification. Based on the results of the three analytical platforms, it was concluded that the major difference in chemical composition of pine species was between species rather than the collection period. Nonetheless, the chemical profiles of resins from different species and collection periods can be well discriminated and correlated to mono- and sesquiterpenes in the case of species and diterpenes for the collection periods.