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A major shoot-specific monoterpene indole alkaloid produced by Psychotria brachyceras, brachycerine, is regulated by either wounding or jasmonate application. Highest concentrations of the alkaloid are found in inflorescences, suggesting a defence role. Brachycerine has antimutagenic and antioxidant properties, capable of quenching singlet oxygen, hydroxyl radical, and superoxide. This study aimed at characterizing the putative role of brachycerine in P. brachyceras responses to wounding and herbivory. Damage to leaves increased the content of brachycerine locally. Wounding did not affect phenolics content in P brachyceras leaves, and no tannins were detected in the species. In generalist herbivore bioassays, neither brachycerine nor P. brachyceras extracts showed toxic effects. In vivo hydrogen peroxide staining assay showed less wound-generated peroxide accumulation in alkaloid treated tissues. This pattern was confirmed in quantitative assays measuring tissue hydrogen peroxide concentrations. Data indicate that brachycerine is not a herbivore deterrent, but rather an indirect chemical defence, modulating oxidative stress caused by mechanical damage.

Monoterpene indole alkaloids (MIAs) encompass plant natural products with important pharmacological relevance. They include the anti-tumoral MIAs found in Catharanthus roseus and Camptotheca acuminata. The often low yields of bioactive alkaloids in plants has prompted research to identify the factors regulating MIA production. Oxidative stress is a general response associated with biotic and abiotic stresses leading to several secondary responses, including elicitation of MIA production. These changes in secondary metabolism may take place directly or via second messengers, such as Ca(2+) and reactive oxygen species (ROS). H2O2 is the main ROS that participates in MIA biosynthesis. This review analyzes the links between oxidative stress, elicitation of bioactive MIA production and their potential roles in antioxidant defense, as well as exploring the implications to developing biotechnological strategies relevant for alkaloid supply.

Psychotria brachyceras Muell. Arg. produces the antioxidant monoterpene indole alkaloid (MIA) brachycerine, which, besides retaining a glucose residue, has its terpenoid moiety derived not from secologanin, but probably from epiloganin, representing a new subclass of MIAs. In this work we showed that osmotic stress agents, such as sodium chloride, sorbitol and polyethylene glycol (PEG), induced brachycerine accumulation in leaf disks of P. brachyceras. Other oxidative stress inducers, such as exposure to aluminum and silver, also increased brachycerine content. Abscisic acid (ABA) treatment was shown to increase brachycerine yield, suggesting its involvement in brachycerine induction during osmotic stress. Ascorbate peroxidase activity was induced in PEG-treated leaf disks, whereas superoxide dismutase (SOD) activity remained unaltered. Assays with specific inhibitors of the cytosolic mevalonate (MVA) and plastidic 2-C-methyl-D-erythritol 4-phosphate (MEP) pathways showed that the terpenoid moiety of brachycerine derived predominantly from the MEP pathway. These results suggest a potential involvement of brachycerine in plant defense against osmotic/oxidative stress damage, possibly contributing to detoxification of hydroxyl radical and superoxide anion as a SOD-like molecule., (Copyright © 2013 Elsevier Masson SAS. All rights reserved.)

UV-B radiation can damage biomolecules, such as DNA, RNA, and proteins, halting essential cellular processes; this damage is partly due to ROS generation. Plant secondary metabolites may protect against UV-B. Psychotria brachyceras Müll. Arg. (Rubiaceae), a subtropical shrub, produces brachycerine, a monoterpene indole alkaloid mainly accumulated in leaf tissues, which displays antioxidant and antimutagenic activities. Exposure of P. brachyceras cuttings to UV-B radiation significantly increases leaf brachycerine concentration. It has been suggested that this alkaloid might contribute to protection against UV-B damage both through its quenching activity on ROS and as UV shield. To identify differentially expressed genes of P. brachyceras in response to UV-B and investigate a possible influence of this stimulus on putative brachycerine-related genes, suppressive subtractive hybridization was applied. Complementary DNA from UV-B-treated leaves for 24 h was used as tester, and cDNA from untreated leaves, as driver. After BLASTX alignments, 134 sequences matched plant genes. Using quantitative RT-PCR, selected genes potentially related to brachycerine showed significant increases in transcription after UV-B exposure: tryptophan decarboxylase, ACC oxidase, UDP-glucose glucosyltransferase, lipase, and serine/threonine kinase. Results suggest a possible involvement of brachycerine in acute UV-B responses and show that alkaloid accumulation seems at least partly regulated at transcriptional level.

The monoterpene indole alkaloid brachycerine from Psychotria brachyceras has been shown to be induced by UV and to have in vitro antioxidant activity, indicating a possible protective role against the secondary effects of this radiation. In this work, we have studied the antioxidant properties of brachycerine and a crude foliar extract from P. brachyceras by using Saccharomyces cerevisiae strains proficient and deficient in antioxidant defenses. The mutagenic and antimutagenic potential of these substances were assayed in S.cerevisiae N123 strain in the presence and absence of H2O2. In addition, we tested the antioxidant capacity of brachycerine and a crude foliar extract from P. brachyceras on hydroxyl radicals (OH) using the hypoxanthine/xanthine oxidase assay. The results show that brachycerine and the crude foliar extract of P. brachyceras have antioxidant and antimutagenic effects in yeast and probably this action is mainly due to the scavenging of OH radicals., ((Copyright) 2007 Wiley-Liss, Inc.)

Monoterpene indole alkaloids (MIAs) are a large class of plant alkaloids with significant pharmacological interest. The sustained production of MIAs at high yields is an important goal in biotechnology. Intensive effort has been expended toward the isolation, cloning, characterization and transgenic modulation of genes involved in MIA biosynthesis and in the control of the expression of these biosynthesis-related genes. At the same time, considerable progress has been made in the detailed description of the subcellular-, cellular-, tissue- and organ-specific expressions of portions of the biosynthetic pathways leading to the production of MIAs, revealing a complex picture of the transport of biosynthetic intermediates among membrane compartments, cells and tissues. The identification of the particular environmental and ontogenetic requirements for maximum alkaloid yield in MIA-producing plants has been useful in improving the supply of bioactive molecules. The search for new bioactive MIAs, particularly in tropical and subtropical regions, is continuously increasing the arsenal for therapeutic, industrially and agriculturally useful molecules. In this review we focus on recent progress in the production of MIAs in transgenic cell cultures and organs (with emphasis on Catharanthus roseus and Rauvolfia serpentina alkaloids), advances in the understanding of in planta spatial-temporal expression of MIA metabolic pathways, and on the identification of factors capable of modulating bioactive alkaloid accumulation in nontransgenic differentiated cultures and plants (with emphasis on new MIAs from Psychotria species). The combined use of metabolic engineering and physiological modulation in transgenic and wild-type plants, although not fully exploited to date, is likely to provide the sustainable and rational supply of bioactive MIAs needed for human well being.

Brachycerine is a monoterpenoid indole alkaloid accumulated in Psychotria brachyceras plants (Rubiaceae). To better understand the accumulation patterns of this alkaloid, we investigated its content in different plant organs from field-grown trees, throughout the seasons, during seedling development, and in response to potential biotic factors regulating its biosynthesis. Quantification by RP-HPLC showed that aerial vegetative organs (green stems, young and old leaves) yielded similar amounts of brachycerine [0.1-0.2% dry weight (DW)]. Brachycerine was not detected in roots. In reproductive structures, the highest brachycerine amounts (0.3% DW) were found in inflorescences. Alkaloid concentration decreased in mature fruits (0.045% DW). The lowest concentration in reproductive organs was observed in quiescent seeds (0.004% DW). Apparently, brachycerine content dropped during radicle emission in germinating seeds. During seedling development, an increase in leaf content from 0.02 to 0.1% DW was observed between the stages of 2 and 14 leaves, respectively. Salicylic acid did not affect brachycerine content. A doubling of alkaloid content was observed in wounded plants, and a threefold induction occurred with jasmonic acid treatment, suggesting that brachycerine biosynthesis is regulated by jasmonate production.

From leaves of Psychotria leiocarpa, an indole alkaloid was isolated to which the structure N,beta-D-glucopyranosyl vincosamide (1) was assigned. This represents the first report of an N-glycosylated monoterpenoid indole alkaloid. In field-grown plants highest amounts of 1 were found in the leaves (2.5% of dry wt) and fruit pulp (1.5% dry wt). Lower amounts were found in the stems (0.2% dry wt) and the seeds (0.1% of dry wt), whereas the alkaloid was not detected in the roots. The accumulation of 1 in aseptic seedlings was also restricted to the shoots and increased with plant age and light exposure, independent of the supply of sucrose in the culture medium.

The effects of ultraviolet (UV) radiation on chlorophyll content and accumulation of the anti-inflammatory monoterpene-indole alkaloid brachycerine in plants and calli of Psychotria brachyceras (Rubiaceae) were investigated. In this study, we also investigated a protective role for brachycerine against stress conditions. Calli and tip cuttings incubated in nutrient media were daily supplemented with 4 or 16 h of UV. High-performance liquid chromatography analyses of methanolic extracts showed only traces of brachycerine in irradiated aseptic cultures, with no alkaloid being observed in control calli. In cuttings, a 10-fold increase in brachycerine content was seen after exposure for 16 h to UV-C, whereas a 4 h daily supplementation doubled the amount of the alkaloid in leaves. Exposure to a UV-B source also doubled the alkaloid yield. In vitro brachycerine was able to quench singlet oxygen. The data indicate a potential protective role for brachycerine against UV radiation, acting as a UV filter (absorption peaks are within the UV range) and a reactive oxygen species scavenger. In addition, UV radiation may be used to increase yields of this compound of pharmaceutical interest.

Brachycerine (1), an unusual alkaloid from the leaves of Psychotria brachyceras, was characterized through spectroscopic data interpretation and its stereochemistry established by NOE difference techniques. Brachycerine (1) was found to be restricted to shoots in rooted cuttings of P. brachyceras (0.018 +/- 0.004% dry weight), and accumulation was unaffected by root induction treatment with auxin.