Metabolic alterations and distribution of five-carbon precursors in jasmonic acid-elicited Catharanthus roseus cell suspension cultures

Plant science

HPLC-DAD DETERMINATION OF BERBERINE, CHELERYTHRINE AND SANGUINARINE IN THE MEXICAN PRICKLY POPPY (Argemone mexicana L. PAPAVERACEAE), A MEDICINAL PLANT

A sensitive, simple, rapid and reliable HPLC-DAD method for the analysis of the benzylisoquinoline alkaloids (BIA's) content in Argemone mexicana (Papaveraceae) is presented. This method allows the simultaneous separation and quantitation of berberine (Bn), chelerythrine (C) and sanguinarine (S) in extracts from A. mexicana tissues, reducing time of analysis in comparison to previous reports. Alkaloids were separated on a C18 Hypersil Gold column using an acetonitrile gradient (20 to 70%) in 1% acetic acid in water. Alkaloids were identified based on retention times and UV spectra and quantified at 254 nm. Linearity between 0.5-20 µg mL-1 was observed for Bn, C and S, with limits of detection (LOD) and quantitation (LOQ) of 0.11 and 0.33 for Bn, 0.10 and 0.30 for C and 0.05 and 0.15 for S, respectively. Maximal intra- and inter-day variation values were < 0.49% in all cases, with alkaloids' recoveries higher than 95%. System suitability tests (SST), including resolution (Rs), retention factor (K'), selectivity (α), tailing factor and number of theoretical plates were performed according to the United States Pharmacopeia (USP), fulfilling recommended values. The method proved to be efficient and reproducible when analyzing different tissues of field-collected A. mexicana plants

Nitrate Promotes Capsaicin Accumulation in Capsicum chinense Immobilized Placentas

In chili pepper’s pods, placental tissue is responsible for the synthesis of capsaicinoids (CAPs), the compounds behind their typical hot flavor or pungency, which are synthesized from phenylalanine and branched amino acids. Placental tissue sections from Habanero peppers (Capsicum chinense Jacq.) were immobilized in a calcium alginate matrix and cultured in vitro, either continuously for 28 days or during two 14-day subculture periods. Immobilized placental tissue remained viable and metabolically active for up to 21 days, indicating its ability to interact with media components. CAPs contents abruptly decreased during the first 7 days in culture, probably due to structural damage to the placenta as revealed by scanning electron microcopy. CAPs levels remained low throughout the entire culture period, even though a slight recovery was noted in subcultured placentas. However, doubling the medium’s nitrate content (from 40 to 80 mM) resulted in an important increment, reaching values similar to those of intact pod’s placentas. These data suggest that isolated pepper placentas cultured in vitro remain metabolically active and are capable of metabolizing inorganic nitrogen sources, first into amino acids and, then, channeling them to CAP synthesis

HPLC-DAD DETERMINATION OF BERBERINE, CHELERYTHRINE AND SANGUINARINE IN THE MEXICAN PRICKLY POPPY (Argemone mexicana L. PAPAVERACEAE), A MEDICINAL PLANT

<div><p>A sensitive, simple, rapid and reliable HPLC-DAD method for the analysis of the benzylisoquinoline alkaloids (BIA's) content in Argemone mexicana (Papaveraceae) is presented. This method allows the simultaneous separation and quantitation of berberine (Bn), chelerythrine (C) and sanguinarine (S) in extracts from A. mexicana tissues, reducing time of analysis in comparison to previous reports. Alkaloids were separated on a C18 Hypersil Gold column using an acetonitrile gradient (20 to 70%) in 1% acetic acid in water. Alkaloids were identified based on retention times and UV spectra and quantified at 254 nm. Linearity between 0.5-20 µg mL-1 was observed for Bn, C and S, with limits of detection (LOD) and quantitation (LOQ) of 0.11 and 0.33 for Bn, 0.10 and 0.30 for C and 0.05 and 0.15 for S, respectively. Maximal intra- and inter-day variation values were < 0.49% in all cases, with alkaloids' recoveries higher than 95%. System suitability tests (SST), including resolution (Rs), retention factor (K'), selectivity (α), tailing factor and number of theoretical plates were performed according to the United States Pharmacopeia (USP), fulfilling recommended values. The method proved to be efficient and reproducible when analyzing different tissues of field-collected A. mexicana plants.</p></div

Folivory elicits a strong defense reaction in Catharanthus roseus: metabolomic and transcriptomic analyses reveal distinct local and systemic responses

Plants deploy distinct secondary metabolisms to cope with environment pressure and to face bioaggressors notably through the production of biologically active alkaloids. This metabolism-type is particularly elaborated in Catharanthus roseus that synthesizes more than a hundred different monoterpene indole alkaloids (MIAs). While the characterization of their biosynthetic pathway now reaches completion, still little is known about the role of MIAs during biotic attacks. As a consequence, we developed a new plant/herbivore interaction system by challenging C. roseus leaves with Manduca sexta larvae. Transcriptomic and metabolic analyses demonstrated that C. roseus respond to folivory by both local and systemic processes relying on the activation of specific gene sets and biosynthesis of distinct MIAs following jasmonate production. While a huge local accumulation of strictosidine was monitored in attacked leaves that could repel caterpillars through its protein reticulation properties, newly developed leaves displayed an increased biosynthesis of the toxic strictosidine-derived MIAs, vindoline and catharanthine, produced by up-regulation of MIA biosynthetic genes. In this context, leaf consumption resulted in a rapid death of caterpillars that could be linked to the MIA dimerization observed in intestinal tracts. Furthermore, this study also highlights the overall transcriptomic control of the plant defense processes occurring during herbivory