Carnivory on demand: phosphorus deficiency induces glandular leaves in the African liana Triphyophyllum peltatum.

Triphyophyllum peltatum, a rare tropical African liana, is unique in its facultative carnivory. The trigger for carnivory is yet unknown, mainly because the plant is difficult to propagate and cultivate. This study aimed at identifying the conditions that result in the formation of carnivorous leaves. In vitro shoots were subjected to abiotic stressors in general and deficiencies of the major nutrients nitrogen, potassium and phosphorus in particular, to trigger carnivorous leaves' development. Adventitious root formation was improved to allow verification of the trigger in glasshouse-grown plants. Among all the stressors tested, only under phosphorus deficiency, the formation of carnivorous leaves was observed. These glandular leaves fully resembled those found under natural growing conditions including the secretion of sticky liquid by mature capture organs. To generate plants for glasshouse experiments, a pulse of 55.4 µM α-naphthaleneacetic acid was essential to achieve 90% in vitro rooting. This plant material facilitated the confirmation of phosphorus starvation to be essential and sufficient for carnivory induction, also under ex vitro conditions. Having established the cultivation of T. peltatum and the induction of carnivory, future gene expression profiles from phosphorus starvation-induced leaves will provide important insight to the molecular mechanism of carnivory on demand.

Naphthylisoquinoline alkaloids and their synthetic analogs as potent novel inhibitors against Babesia canis in vitro.

Babesia canis is the predominant and clinically relevant canine Babesia species in Europe. Transmitted by vector ticks, the parasite enters red blood cells and induces a severe, potentially fatal hemolytic anemia. Here, we report on the antibabesial activities of three extracts of the West African tropical plant species Triphyophyllum peltatum (Dioncophyllaceae) and Ancistrocladus abbreviatus (Ancistrocladaceae) and of 13 genuine naphthylisoquinoline alkaloids isolated thereof. Two of the extracts and eight of the alkaloids were found to display strong activities against Babesia canis in vitro. Among the most potent compounds were the C,C-coupled dioncophyllines A (1a) and C (2) and the N,C-linked alkaloids ancistrocladium A (3) and B (4), with half-maximum inhibition concentration (IC50) values of 0.48 µM for 1a, 0.85 µM for 2, 1.90 µM for 3, and 1.23 µM for 4. Structure-activity relationship (SAR) studies on a small library of related genuine analogs and non-natural synthetic derivatives of 1a and 2 revealed the likewise naturally occurring alkaloid N-methyl-7-epi-dioncophylline A (6b) to be the most potent (IC50, 0.14 µM) among the investigated compounds. Although none of the tested naphthylisoquinolines showed 100 % inhibition of parasite infection - as displayed by imidocarb dipropionate (IC50, 0.07 µM), which was used as a positive control - the antibabesial potential of the dioncophyllines A (1a) and C (2) and related compounds such as 6b, its atropo-diastereomer 6a (IC50, 1.45 µM), and 8-O-(p-nitrobenzyl)dioncophylline A (14) (IC50, 0.82 µM) is to be considered as high. The SAR results showed that N-methylation and axial chirality exert a strong impact on the antibabasial activities of the naphthylisoquinolines presented here, whereas dimerization, as in jozimine A2 (5) (IC50, 140 µM), leads to a significant decrease of activity against B. canis. Alkaloids displaying good to high activities against B. canis like the dioncophyllines 1a, 2, 6a, and 6b were found to cause only a small degree of hemolysis (< 0.7 %), whereas compounds with moderate to weak antibabesial activities such as 6-O-methyl-4'-O-demethylancistrocladine (15a) (IC50, 14.0 µM) and its atropo-diastereomer 6-O-methyl-4'-O-demethylhamatine (15b) (IC50, 830 µM) caused a high degree of hemolysis (7.3 % for 15a and 11.2 % for 15b). In this respect, the most effective anti-Babesia naphthylisoquinolines are also the safest ones.

The Plant-Derived Naphthoquinone Droserone Inhibits In Vitro Measles Virus Infection.

The naphthoquinone droserone (1) is a natural product occurring in dicotyledonous plants. We have now observed that the addition of 1 during infection of tissue culture cells with measles virus considerably reduced the infection. Interestingly, the infection was inhibited only when droserone (1) was added during virus entry, but not when added to the cells prior to virus uptake or after virus uptake. These findings suggest that 1 interacts with viral particles to reduce infectivity. The formation of progeny measles virus particles was inhibited to 50 % by droserone (1) at a concentration (IC50) of approximately 2 µM with a half-maximal cytotoxicity (CC50) of about 60 µM for Vero cells. Other tested naphthoquinone derivatives, among them the likewise natural plumbagin (2), but also synthetic analogs, were either more cytotoxic or not as effective as 1. Thus, our data do not support the development of naphthoquinone derivatives into antiviral compounds, but suggest that they may be interesting research tools to study measles virus entry into cells.

Axially Chiral Dimeric Naphthalene and Naphthoquinone Metabolites, from Root Cultures of the West African Liana Triphyophyllum peltatum.

Root cultures of the West African liana Triphyophyllum peltatum were initiated from stem explants of in vitro cultivated shoots. From these organ cultures, three new binaphthalenes, one binaphthoquinone, and two (bi)naphthalene glucosides were isolated, with substitution patterns related to those of the naphthylisoquinoline alkaloids, which are the "normal" main metabolites of T. peltatum. The structures of the diglucoside dioncoquinoside A (1) and of the axially chiral biaryls triphyoquinols A1 (3), A2 (4), and B (5), triphyoquinoside A (6), and triphyoquinone A (7) were elucidated by spectroscopic analysis (HRESIMS, 1D and 2D NMR) and by application of electronic circular dichroism (ECD) spectroscopy in combination with the exciton chirality method and quantum-chemical ECD calculations. The root cultures likewise produced the known alkaloids dioncophylline A (8), 5'-O-demethyldioncophylline A (9), dioncopeltine A (10), habropetaline A (11), and 5'-O-methyldioncophylline D (12a/b), the naphthalene glucoside plumbaside A (2), and the naphthoquinones plumbagin (13), droserone (14), and 8-hydroxydroserone (15).

Naphthylisoquinoline alkaloids potential drug leads.

Naphthylisoquinolines are a group of structurally diverse secondary metabolites, consisting of naphthalene and isoquinoline moieties. Naturally occurring naphthylisoquinolines have so far been found only in the small palaeotropic families Dioncophyllaceae and Ancistrocladaceae. They have been shown to exhibit a diverse array of biological activities. Herein, we review the research on the occurrence, isolation, identification, biological activities, and biosynthesis of this class of compounds published from 1995 till now. Moreover, their chemotaxonomic relevance and molecular targets of action have been discussed. More than 125 metabolites are described and 99 references are cited.

Anti-tumoral activities of dioncoquinones B and C and related naphthoquinones gained from total synthesis or isolation from plants.

Dioncoquinones belong to a family of natural naphthoquinone products of interest due to their promising anti-tumoral and anti-infective activities. In particular, dioncoquinones A (5) and B (6) have been shown to be highly active against Leishmania major and multiple myeloma cells without any significant toxicity toward normal blood cells. Their effective concentrations against multiple myeloma cell lines were similar to those of melphalan, a well known DNA-alkylating agent used in a standard therapy against B cell lymphoma and multiple myeloma. We report on the first total synthesis of the highly oxygenated anti-tumoral agent dioncoquinone B (6) and the isolation of its new, even higher-oxygenated analogs, dioncoquinones C (7), D (8), and E (9), from cell cultures of Triphyophyllum peltatum. In addition, several derivatives of these compounds were synthesized, including dioncoquinone C (7), and a small library of naphthoquinones was created. Furthermore, the first structure-activity relationship (SAR) study on this class of compounds was conducted, showing that each of the three hydroxy groups, at C-3, C-5, and C-6, is required for improved anti-tumoral activities and decreased cytotoxicities.

Antitumoral and antileishmanial dioncoquinones and ancistroquinones from cell cultures of Triphyophyllum peltatum (Dioncophyllaceae) and Ancistrocladus abbreviatus (Ancistrocladaceae).

From the methanolic extracts of solid callus cultures from two species of the closely related palaeotropical plant families Dioncophyllaceae and Ancistrocladaceae seven new natural naphthoquinones were isolated, dioncoquinones A (4) and B (5) from Triphyophyllum peltatum, and ancistroquinones B (6), C (7), D (9), E (10), and F (12) from Ancistrocladus abbreviatus. Their structures were elucidated by spectroscopic, chemical, and computational methods. Furthermore, the already known naphthoquinones plumbagin (2), droserone (3), malvone A (8), and nepenthone A (11) were found in the extract of A. abbreviatus. Dioncoquinones A (4) and B (5) showed good - and specific - activity against Leishmania major, while they were not active against other protozoic parasites. Moreover, treatment with 4 and 5 strongly induced apoptosis in human tumor cells derived from two different B cell malignancies, B cell lymphoma and multiple myeloma, without any significant toxicity towards normal peripheral mononuclear blood cells.

Ultrasensitive in situ tracing of the alkaloid dioncophylline A in the tropical liana Triphyophyllum peltatum by applying deep-UV resonance Raman microscopy.

UV resonance Raman microspectroscopy was applied for a localization of the antiplasmodial naphthylisoquinoline alkaloid dioncophylline A in very low concentrations in different parts of the samples (e.g., in the roots) of the tropical liana Triphyophyllum peltatum. The application of resonance Raman microspectroscopy was characterized by a very high sensitivity and selectivity. It was possible to assign the resonance Raman spectra of dioncophylline A, dioncophylline C, and dioncopeltine A by means of a combination of NIR Raman spectroscopy and DFT calculations. The UV resonance Raman spectra of T. peltatum are very well resembled by the spectra of dioncophylline A, while they can be clearly distinguished from the spectra of dioncophylline C and dioncopeltine A. This distinction between the various naphthylisoquinolines was possible by the two modes at 1356 and 1613 cm-1. These two modes were assigned to C=C stretching and CH bending vibrations. The presented results of a highly sensitive and selective in situ localization of the active agent dioncophylline A in different parts of the plant material of T. peltatum are of high importance for the acquisition of new antimalarials and for plant science in general.

In vivo localization and identification of the antiplasmodial alkaloid dioncophylline A in the tropical liana Triphyophyllum peltatum by a combination of fluorescence, near infrared Fourier transform Raman microscopy, and density functional theory calculations.

Near infrared Fourier transform (NIR FT) micro Raman spectroscopy in combination with density functional theory (DFT) calculations has been applied for an in vivo localization of the antiplasmodial naphthylisoquinoline alkaloid dioncophylline A (1) in the tropical liana Triphyophyllum peltatum. Fluorescence microscopy images suggest finding this active agent in 10 mum big inclusions located in the cortex of the stem or the beginning of the leaves. By means of spatially resolved FT Raman micro spectroscopy, we could detect dioncophylline A (1) in these inclusions. FT Raman spectroscopy is an extremely selective tool capable of differentiating between various structurally similar naphthylisoquinoline alkaloids. With the help of DFT calculations, we succeeded in assigning the differences found in the FT Raman spectra of the various naphthylisoquinolines to nuC=C vibrations of the naphthyl ring. The presented results are of relevance for the investigation and extraction of new antimalarial active agents.