Method for determination of pyrrolizidine alkaloids and their metabolites by high-performance liquid chromatography

An improved method utilizing reverse-phase liquid chromatography on a styrene-divinylbenzene column (PRP-1) and ultraviolet detection was developed for the simultaneous determination of the pyrrolizidine alkaloids (PAs) senecionine, seneciphylline, and retrorsine and their major metabolites produced during in vitro transformation of PAs by microsomal enzyme systems. The procedure employs direct injection of the 46,000g supernatant of the microsomal reaction mixture directly onto the column, and elution with a 0.1 M NH4OH-acetonitrile gradient. The method is very gentle, simple, and fast with excellent precision since no prior extraction, preconcentration, or derivatization steps are required. Using this procedure 6,7-dihydro-7-hydroxy-1-hydroxymethyl-5H-pyrrolizine and the corresponding PA N-oxides were shown to be the major microsomal metabolites of the PAs examined. The detection limit of these metabolites was approximately 1 microM.     

Seasonal variability in pyrrolizidine alkaloids in Senecio inaequidens from the Val Venosta (Northern Italy)

Senecio inaequidens is a neophyte originating from South Africa that has managed to spread to Europe and colonize large areas. This plant has toxic pyrrolizidine alkaloids (PAs) that represent a health risk to humans and animals. The aim of this work was to measure the content of PAs of these plants in continuous intervals over the entire growing season. The plants were separated into their organs, such as sprouts, stems, leaves and inflorescences. PAs were extracted from the dried plants using an acidic methanol solution, purified and measured by GC/MS. The average PA content of plants over the growing season was 0.33% of dry weight. The highest PA levels were about 1% in the dry weight and were found in the young sprouts and flower heads. There were nine PAs present, of which six could be identified. The main alkaloid was retrorsine followed by senecivernine, senecionine, integerrimine, usaramine and seneciphylline. Due to the high PA content of the inflorescences, the long flowering period and the rapid invasion dynamics, this species presents a high health risk for humans and animals.     

Modulation of gastrin and epidermal growth factor by pyrrolizidine alkaloids obtained from Senecio brasiliensis in acute and chronic induced gastric ulcers

We investigated the antiulcerogenic activity of pyrrolizidine alkaloids (PAs) integerrimine, retrorsine, senecionine, usaramine and seneciplhylline, an alkaloidal extract obtained from Senecio brasiliensis. The PA extract demonstrated significantly activity in both, acute and chronic gastric ulcers on rats. The effects of PA extract were dose dependent. The mechanisms implicated on this activity were evaluated by determination of gastrin plasma levels in rats subjected to the acute treatment with PA extract and by expression of mRNA of Epidermal Growth Factor (EGF) after chronic treatment with this extract. The results showed that the PA extract increased both the levels of gastrin and the expression of EGF on these animals. Moreover, the histological examinations showed a reduction of exfoliation of superficial cells, hemorrhages and blood cell infiltration. We concluded that the PAs showed an important and qualitative antiulcerogenic activity mediated by increase in gastrin secretion and mRNA expression of EGF.     

Frequent gain and loss of pyrrolizidine alkaloids in the evolution of Senecio section Jacobaea (Asteraceae)

Pyrrolizidine alkaloids (PAs) of the macrocyclic senecionine type are secondary metabolites characteristic for most species of the genus Senecio (Asteraceae). These PAs are deterrent and toxic to most vertebrates and insects and provide plants with a chemical defense against herbivores. We studied the PA composition of 24 out of 26 species of Senecio section Jacobaea using GC-MS. The PA profiles of eight of these species have not been studied before and additional PAs were identified for most other species that were included in previous studies. With one exception (senecivernine) all 26 PAs identified in sect. Jacobaea can be regarded as derivatives of the biosynthetic backbone structure senecionine. Based on the PA profiles of the species of sect. Jacobaea and the results of previous tracer studies, we constructed two hypothetical biosynthetic scenarios of senecionine diversification. Both scenarios contain two major reactions: the conversion of the necine base moiety retronecine into the otonecine moiety and site-specific epoxidations within the necic acid moiety. Further reactions are site-specific hydroxylations, sometimes followed by O-acetylations, site-specific dehydrogenations, E, Z-isomerizations, and epoxide hydrolysis and chlorolysis. The GC-MS data and both biosynthetic scenarios were subsequently used to study the evolution of PA formation in sect. Jacobaea by reconstructing the evolutionary history of qualitative PA variation in this section. This was achieved by optimizing additive presence/absence data of PAs and types of enzymatic conversions on a maximum parsimony cladogram of section Jacobaea inferred from DNA sequence and morphological data. Besides showing large intra- and interspecific variation, PA distribution appears to be largely incidental within the whole clade. These results together with the finding that all but one of the PAs identified in sect. Jacobaea are also present in species of other sections of Senecio indicate that differences in PA profiles in Senecio can not be explained by the gain and loss of PA specific genes, but rather by a transient switch-off and switch-on of the expression of genes encoding PA pathway-specific enzymes.     

Concomitant occurrence of pyrrolizidine and quinolizidine alkaloids in the hemiparasite Osyris alba L. (Santalaceae)

The investigation of the alkaloid extracts of the hemiparasitic plant Osyris alba, collected from three different localities in southern France, revealed the concomitant presence of both pyrrolizidine (PA) and quinolizidine (QA) alkaloids in the samples from two of these localities. The sample from the third locality contained only PAs. The eight QAs identified were sparteine, N-methylcytisine, cytisine, methyl-12-cytisine acetate, hydroxy-N-methylcytisine, N-acetylcytisine, lupanine, and anagyrine. Of the eleven detected PAs, eight were identified as chysin A, chysin B, 1-carboxypyrrolizidine-7-olide, senecionine, integerrimine, retrorsine, senecivernine and a new alkaloid janfestine (7R-hydroxychysin A or 1R-carbomethoxy-7R-hydroxypyrrolizidine). PAs were mainly present as their N-oxides This is, to our knowledge, the first report demonstrating the simultaneous presence of two classes of alkaloids, quinolizidine and pyrrolizidine alkaloids, in a single parasitic plant. As these alkaloids do not occur in the same host plant, the results indicate that Osyris must have tapped more than one host plant concomitantly. Since both quinolizidine and pyrrolizidine alkaloids serve as defence compounds against herbivores, affecting different molecular targets, the simultaneous acquisition of the two types of alkaloids by a single plant could provide a novel mode of defence of hemiparasites against herbivores.     

Root damage and aboveground herbivory change concentration and composition of pyrrolizidine alkaloids of Senecio jacobaea

Thus far not many studies focussed on how herbivory in one plant part affects plant defence in the other. The effects of root damage and a leaf-feeding herbivore (Mamestra brassicae) on pyrrolizidine alkaloid (PA) levels of Senecio jacobaea were investigated in a controlled environment. Three cloned S. jacobaea genotypes, which differed in PA concentrations, received four treatments: (1) no damage, (2) root damage (removing half of the root system), (3) shoot herbivory by M. brassicae larvae, (4) root damage and shoot herbivory.Shoot herbivory did not significantly affect shoot biomass, while root damage decreased both root and shoot biomass. Shoot herbivory decreased PA concentrations in the roots. Conversely, root damage increased PA concentrations in the roots. Alkaloid concentrations in the shoot showed a weak response to root damage, shoot herbivory had no effect on PA levels in the shoot. The effect of damage on the allocation of PAs to shoot and roots depended on genotype. One genotype allocated more PAs to the damaged site, another genotype did not change allocation and the third genotype allocated more PAs to the shoot if the roots were damaged. Changes in PA composition were observed in one genotype. Shoot herbivory increased erucifoline concentrations in the shoot and decreased concentrations of senecionine in the roots. In conclusion, we have shown that even in an alleged constitutively defended plant, damage of one compartment affects secondary metabolite level in the other.     

The genotype dependent presence of pyrrolizidine alkaloids as tertiary amine in Jacobaea vulgaris

Secondary metabolites such as pyrrolizidine alkaloids (PAs) play a crucial part in plant defense. PAs can occur in plants in two forms: tertiary amine (free base) and N-oxide. PA extraction and detection are of great importance for the understanding of the role of PAs as plant defense compounds, as the tertiary PA form is known for its stronger influence on several generalist insects, whereas the N-oxide form is claimed to be less deterrent. We measured PA N-oxides and their reduced tertiary amines by liquid chromatography-tandem mass spectrometry (LC-MS/MS). We show that the occurrence of tertiary PAs is not an artifact of the extraction and detection method. We found up to 50% of tertiary PAs in shoots of Jacobine - chemotype plants of Jacobaea vulgaris. Jacobine and its derivatives (jacoline, jaconine, jacozine and dehydrojaconine) may occur for more than 20% in reduced form in the shoots and more than 10% in the roots. For 22 PAs detected in F(2) hybrids (J. vulgaris × Jacobaea aquatica), we calculate the tertiary amine percentage (TA%=the tertiary amine concentration/(tertiary amine concentration+the corresponding N-oxide concentration) × 100). We found that the TA% for various PAs was genotype-dependent. Furthermore, TA% for the different PAs were correlated and the highest correlations occurred between PAs which share high structural similarity.     

Feeding responses of specialist herbivores to plant extracts and pure allelochemicals: effects of prolonged exposure

We investigated the possibility of decreased feeding deterrent response following prolonged exposure in three specialist herbivores, Plutella xylostella (larvae), Pseudaletia unipuncta (larvae), and Epilachna varivestis (adults) through leaf disc choice and no‐choice tests. Neonate larvae (<24 h old) of the three species were reared on their host plants sprayed with the antifeedants until tested. Our results demonstrated that Ps. unipuncta did not show a decrease in feeding deterrent response to extracts of Melia volkensii (choice and no‐choice tests) or oregano (Origanum vulgare) (choice test) following prolonged exposure. Plutella xylostella exhibited a decrease in feeding deterrent response to M. volkensii in a choice test only, but not to oregano. Although not significant, we did find a trend toward decreasing feeding deterrent response to M. volkensii by both species in no‐choice tests. However, both species exhibited a significant decrease in feeding deterrent response to pure allelochemicals (Ps. unipuncta to thymol, and P. xylostella to thymol and toosendanin) compared with the naïve groups, following prolonged exposure in leaf disc choice tests. Epilachna varivestis showed a significant decrease in feeding deterrent responses to both oregano and thymol in leaf disc choice tests. We conclude from our present and previous studies that not only are there interspecific differences between generalist and specialist species but also among specialist species.     

Variation in pyrrolizidine alkaloid patterns of Senecio jacobaea

We studied the variation in pyrrolizidine alkaloid (PA) patterns of lab-grown vegetative plants of 11 European Senecio jacobaea populations. Plants were classified as jacobine, erucifoline, mixed or senecionine chemotypes based on presence and absence of the PAs jacobine or erucifoline. Due to the presence of jacobine, total PA concentration in jacobine chemotypes was higher than in erucifoline chemotypes. Both relative and absolute concentrations of individual PAs differed between half-sib and clonal families, which showed that variation in PA patterns had a genetic basis. Within most populations relative abundance of PAs varied considerably between individual plants. Most populations consisted either of the jacobine chemotype or of the erucifoline chemotype, sometimes in combination with mixed or senecionine chemotypes.     

Changes in Senecio grisebachii pyrrolizidine alkaloids abundances and profiles as response to soil quality

Abstract

Senecio grisebachii Baker is an invasive weed considered to be toxic due to the presence of pyrrolizidine alkaloids (PA) in its tissues. The PA production by S. grisebachii aerial parts was evaluated in samples grown in two Argentinean Rolling Pampa fields with the same kind of soil but differing in the length of their exploitation period by conventional tillage practices and, consequently, in their deterioration level.

We found significant differences in the relative concentrations of seven alkaloids between samples taken from the two fields. Seneciphylline was the most abundant alkaloid in inflorescences from less deteriorated soil (LD) while senecionine was the major one in those from highly deteriorated soil (D) being followed by seneciphylline, integerrimine, and minor amounts of spartiodine, jacobine, jacozine and retrorsine. A significant increase in total alkaloid content (TAC) was observed in inflorescences from samples growing in D soil (3.52±0.20 mg/g DW) when comparing with those from samples grown in LD one (3.23±0.26).     

Habituation in Frankliniella occidentalis to deterrent plant compounds and their blends

Feeding and oviposition deterrence of three secondary plant compounds and their 1:1 blends to adult female Frankliniella occidentalis Pergande (Thysanoptera: Thripidae) and the potential for habituation of the thrips to the pure compounds and the 1:1 blends at various concentrations were investigated. In choice assays, we tested dose‐dependent feeding and oviposition deterrence of the two fatty acid derivatives methyl jasmonate and cis‐jasmone, the phenylpropanoid allylanisole, and their blends when directly applied to bean leaf discs. The concentration required to reduce the feeding damage by 50% relative to the control treatment (FDC₅₀) was lowest for cis‐jasmone and highest for allylanisole. The feeding deterrent effect of both jasmonates was increased when blended with allylanisole. Feeding deterrence and oviposition deterrence were strongly correlated. In no‐choice assays conducted over four consecutive days, we discovered that dilutions at low concentrations (FDC₁₅) applied to bean leaves resulted in habituation to the deterrents, whereas no habituation occurred at higher concentrations (FDC₅₀). We observed a tendency that the 1:1 blends reduce the probability that thrips habituate to the deterrent compounds. Our results may be useful in the development of integrated crop protection strategies with the implementation of allelochemicals as pest behaviour‐modifying agents.     

Biochemical strategy of sequestration of pyrrolizidine alkaloids by adults and larvae of chrysomelid leaf beetles

Tracer feeding experiments with (14)C-labeled senecionine and senecionine N-oxide were carried out to identify the biochemical mechanisms of pyrrolizidine alkaloid sequestration in the alkaloid-adapted leaf beetle Oreina cacaliae (Chrysomelidae). The taxonomically closely related mint beetle (Chrysolina coerulans) which in its life history never faces pyrrolizidine alkaloids was chosen as a 'biochemically naive' control. In C. coerulans ingestion of the two tracers resulted in a transient occurrence of low levels of radioactivity in the hemolymph (1-5% of radioactivity fed). With both tracers, up to 90% of the radioactivity recovered from the hemolymph was senecionine. This indicates reduction of the alkaloid N-oxide in the gut. Adults and larvae of O. cacaliae sequester ingested senecionine N-oxide almost unchanged in their bodies (up to 95% of sequestered total radioactivity), whereas the tertiary alkaloid is converted into a polar metabolite (up to 90% of total sequestered radioactivity). This polar metabolite, which accumulates in the hemolymph and body, was identified by LC/MS analysis as an alkaloid glycoside, most likely senecionine O-glucoside. The following mechanism of alkaloid sequestration in O. cacaliae is suggested to have developed during the evolutionary adaptation of O. cacaliae to its alkaloid containing host plant: (i) suppression of the gut specific reduction of the alkaloid N-oxides, (ii) efficient uptake of the alkaloid N-oxides, and (iii) detoxification of the tertiary alkaloids by O-glucosylation. The biochemical mechanisms of sequestration of pyrrolizidine alkaloid N-oxides in Chysomelidae leaf beetles and Lepidoptera are compared with respect to toxicity, safe storage and defensive role of the alkaloids.     

Pyrrolizidine alkaloids in the antipodean genus Brachyglottis (Asteraceae)

The phylogeny of the genus Brachyglottis suggests that its constituent species should contain pyrrolizidine alkaloids. Consistent with this hypothesis, and the established occurrence of such alkaloids in Brachyglottis repanda, Brachyglottis kirkii, and Brachyglottis hectori, an investigation of Brachyglottis adamsii revealed the presence of senecionine and retrorsine; Brachyglottis huntii was found to contain senkirkine and retrorsine; 7-O-angelylheliotridine was the predominant alkaloid in Brachyglottis perdicioides, and the same alkaloid together with senecionine, senkirkine and intergerrimine was present in the Brachyglottis hectori × B. perdicioides “Alfred Atkinson” horticultural hybrid; Brachyglottis sciadophila contained clivorine and neopetasitenine (acetylfukinotoxin); the latter alkaloid was also present in B. kirkii together with the previously reported senkirkine and senkirkine 12-acetate.     

Attract and deter: a dual role for pyrrolizidine alkaloids in plant–insect interactions

Pyrrolizidine alkaloids (PAs) are the major defense compounds of plants in the Senecio genus. Here I will review the effects of PAs in Senecio on the preference and performance of specialist and generalist insect herbivores. Specialist herbivores have evolved adaptation to PAs in their host plant. They can use the alkaloids as cue to find their host plant and often they sequester PAs for their own defense against predators. Generalists, on the other hand, can be deterred by PAs. PAs can also affect survival of generalist herbivores. Usually generalist insects avoid feeding on young Senecio leaves, which contain a high concentration of alkaloids. Structurally related PAs can differ in their effects on insect herbivores, some are more toxic than others. The differences in effects of PAs on specialist and generalists could lead to opposing selection on PAs, which may maintain the genetic diversity in PA concentration and composition in Senecio species.     

Chemical diversity and variation of pyrrolizidine alkaloids of the senecionine type: biological need or coincidence?

Laboratory and field tracer experiments with ¹⁴C-labelled senecionine N–oxide (SO) and distant biosynthetic precursors such as [¹⁴C]putrescine revealed that pyrrolizidine alkaloid N-oxides (PAs) in Senecio vernalis Waldstr. & Kit. (Asteraceae) show no significant turnover over periods of up to 29 d. However, PAs are spatially mobile, they are continuously allocated, and labelled PAs are even detectable in leaves and capitula developed weeks after tracer application. Chemical diversification of SO, the common product of PA biosynthesis in roots, was studied in five Senecio species (i.e. S. vernalis Waldstr. & Kit., S. vulgaris L, S. inaequidens DC, two chemotypes of S. jacobaea L. and S. erucifolius L.). Tracer experiments revealed that shoots are capable of transforming [¹⁴ C]SO into the unique species–specific PA patterns. Within a plant, the transformation efficiency of SO can vary quantitatively and qualitatively between shoot organs (i.e. leaves, stems and inflorescences). All transformations proceed position-specifically and stereoselectively. They comprise simple one-step or two-step reactions such as hydroxylations, epoxidations, dehydrogenations, and O-acetylations, as well as the more complex conversion of the retronecine into the otonecine base moiety (e.g. SO into senkirkine). Taking all the evidence together, the qualitative and quantitative composition of the Senecio PA pattern is a dynamic and sensitive equilibrium between a number of interacting processes: (i) constant rate of de-novo synthesis of SO in roots, (ii) continuous long-distance translocation of SO into shoots, (iii) efficiency of SO transformations which may vary between plant organs, (iv) continuous allocation of PAs in the plant, and (v) efficiency and tissue selectivity of vacuolar storage. We suggest that in constitutive plant defence, without significant turnover of its components, such a highly plastic system provides a powerful strategy to successfully defend and possibly escape herbivory. Received: 27 March 1998 / Accepted: 19 May 1998     

The effect of nutrients on pyrrolizidine alkaloids in Senecio plants and their interactions with herbivores and pathogens

The aim of this review is to combine the knowledge of studies on effects of nutrients on pyrrolizidine alkaloids (PAs) in Senecio with those studies of effects of PAs on herbivores and pathogens in order to predict the effects that nutrients may have on herbivores and pathogens via changes in PAs. We discuss whether these predictions match with the outcome of studies where the effect of nutrients on herbivores and insects were measured. PA concentrations in S. jacobaea, S. vulgaris and S. aquaticus were mostly reduced by NPK fertilization, with genotype-specific effects occurring. Plant organs varied in their response to increased fertilization; PA concentrations in flowers remained constant, while shoot and roots were mostly negatively affected. Biomass change is probably largely responsible for the change in concentrations. Nutrients affect both the variety and the levels of PAs in the plant. The reduced PA concentrations after NPK fertilization was expected to benefit herbivores, but no or negative responses from insect herbivores were observed. Apparently other changes in the plant after fertilization are overriding the effect of PAs. Pathogens do seem to benefit from the lower PA concentrations after fertilization; they were more detrimental to fertilized plants than to unfertilized control plants. Future studies should include the effect of each element of nutrients separately and in combinations in order to gain more insight in the effect of specific nutrients on PA content in Senecio plants.     

Selective uptake of pyrrolizidine N-oxides by cell suspension cultures from pyrrolizidine alkaloid producing plants

The N-oxides of pyrrolizidine alkaloids such as senecionine or monocrotaline are rapidly taken up and accumulated by cell suspension cultures obtained from plants known to produce pyrrolizidines, i.e. Senecio vernalis, vulgaris, viscosus (Asteraceae) and Symphytum officinale (Boraginaceae). The transport of the N-oxides into the cells is a specific and selective process. Other alkaloid N-oxides such as sparteine N-oxide are not taken up. Cell cultures from plant species which do not synthesize pyrrolizidine alkaloids are unable to accumulate pyrrolizidine N-oxides. The suitability of the pyrrolizidine N-oxides in alkaloid storage and accumulation is emphasized.     

Production and sex‐pheromonal activity of alkaloid‐derived androconial compounds in the danaine butterfly,Parantica sita (Lepidoptera: Nymphalidae: Danainae)

Close associations of certain lepidopteran taxa with pyrrolizidine alkaloids (PAs), a typical class of plant secondary metabolites, have been well documented from the perspective of evolutionary ecology. Male danaine butterflies are thought to utilize PAs as precursors for the production of dihydropyrrolizines [e.g. danaidone (DO) and hydroxydanaidal (HD)] in their two distinct androconial organs, viz. alar scent organs (sex brands) and abdominal hairpencils. However, little is known about the quantitative profiles of these compounds in danaines, the mechanism for their formation in the androconial organs, or their biological functions, particularly in mating behaviour. The present study addressed these unanswered questions posed for males of the danaine butterfly, Parantica sita. Chemical analyses of androconial extracts revealed considerable seasonal/regional and individual variations of the amounts of DO (the major dihydropyrrolizine produced) and 7R‐HD (the 7R‐enantiomer of HD detected in this study) found in the two organs. These variations seemed to depend primarily on the age of the male and the phenological traits of PA‐containing plants available. Males were found to acquire an adequate capability to produce DO ~1 week after eclosion. DO was shown to be produced exclusively in the sex brand and subsequently physically transferred to the hairpencil through a contact behaviour between the two organs, here termed ‘perfuming behaviour’. The results of behavioural experiments with PA‐fed and PA‐unfed males that were allowed to compete for mates, combined with the positive electroantennographic (EAG) responses of the female, to both DO and 7R‐HD, led to the conclusion that either or both of these compounds can act as the sex pheromone. Oral administration of PAs to males indicated that DO can be biosynthesized from various PA precursors, while 7R‐HD, unlike in arctiid moths, is derived only from PAs with the 7R‐configuration. The putative biosynthetic pathways of DO and 7R‐HD, and the evolutionary provenance of the binate androconial system in the Danainae are also discussed.     

Pyrrolizidine alkaloids of the endemic Mexican genus Pittocaulon and assignment of stereoisomeric 1,2-saturated necine bases

The endemic Mexican genus Pittocaulon (subtribe Tussilagininae, tribe Senecioneae, Asteraceae) belongs to a monophyletic group of genera distributed in Mexico and North America. The five Pittocaulon species represent shrubs with broom-like succulent branches. All species were found to contain pyrrolizidine alkaloids (PAs). With one exception (i.e., stems of Pittocaulon velatum are devoid of PAs) PAs were found in all plant organs with the highest levels (up to 0.3% of dry weight) in the flower heads. Three structural types of PAs were found: (1) macrocyclic otonecine esters, e.g. senkirkine and acetylpetasitenine; (2) macrocyclic retronecine esters, e.g. senecionine, only found in roots, and (3) monoesters of 1,2-saturated necines with angelic acid. For an unambiguous assignment of the different stereoisomeric 1,2-saturated necine bases a GC-MS method was established that allows the separation and identification of the four stereoisomers as their diacetyl or trimethylsilyl derivatives. All otonecine esters that generally do not form N-oxides and the 1,2-saturated PAs were exclusively found as free bases, while the 1,2-unsaturated 7-angeloylheliotridine occurring in P. velatum was found only as its N-oxide. In a comparative study the 1H and 13C NMR spectra of the four stereoisomeric necine bases were completely assigned by the use of DEPT-135, H,H-COSY, H,C-HSQC and H,H-NOESY experiments and by iterative analysis of the 1H NMR spectra. Based on these methods the PA monoesters occurring in Pittocaulon praecox and P. velatum were assigned as 7-O-angeloyl ester respectively 9-O-angeloyl ester of dihydroxyheliotridane which could be identified for the first time as naturally occurring necine base. Unexpectedly, in the monoesters isolated from the three other Pittocaulon species dihydroxyheliotridane is replaced by the necine base turneforcidine with opposite configuration at C-1 and C-7. The species-specific and organ-typical PA profiles of the five Pittocaulon species are discussed in a biogenetic context.     

Site of synthesis,metabolism and translocation of senecionine N-oxide in cultured roots of Senecio erucifolius

Root cultures of Senecio erucifolius (Asteraceae) efficiently took up and incorporated [¹⁴C]putrescine and [¹⁴C]arginine into the pyrrolizidine alkaloid (PA) senecionine N-oxide. Pulse-chase experiments covering a growth period of 10 to 19 days revealed the absence of any significant alkaloid turnover. The only metabolic activity was a slow but progressive transformation of senecionine N-oxide into its dehydrogenation product, seneciphylline N-oxide. Tracer experiments with single roots showed that the sites of enhanced PA synthesis coincided with the sites of preferred protein synthesis, i.e. root apices, indicating a close correlation between growth activity and alkaloid synthesis. Long-term pulse-chase experiments (10 to 12 days) with ¹⁴C-labelled arginine, putrescine and senecionine fed to single roots indicated that in spite of its metabolic inertia, senecionine N-oxide is a mobile compound which is translocated into tissues newly grown during the chase.Dedicated to Dr. Friedrich Constabel on the occasion of his 60th birthday