Pyrrolizidine alkaloids in human diet.

Pyrrolizidine alkaloids are the leading plant toxins associated with disease in humans and animals. Upon ingestion, metabolic activation in liver converts the parent compounds into highly reactive electrophiles capable of reacting with cellular macromolecules forming adducts which may initiate acute or chronic toxicity. The pyrrolizidine alkaloids present a serious health risk to human populations that may be exposed to them through contamination of foodstuffs or when plants containing them are consumed as medicinal herbs. Some pyrrolizidine alkaloids (PA) adducts are persistent in animal tissue and the metabolites may be re-released and cause damage long after the initial period of ingestion. PAs are also known to act as teratogens and abortifacients. Chronic ingestion of plants containing PAs has also led to cancer in experimental animals and metabolites of several PAs have been shown to be mutagenic in the Salmonella typhimurium/mammalian microsome system. However, no clinical association has yet been found between human cancer and exposure to PAs. Based on the extensive reports on the outcome of human exposure available in the literature, we conclude that while humans face the risk of veno-occlusive disease and childhood cirrhosis PAs are not carcinogenic to humans.     

Genetic variation in constitutive and inducible pyrrolizidine alkaloid levels inCynoglossum officinale L.

The constitutive pyrrolizidine alkaloid (PA) concentration of both shoots and roots differed significantly between 17 selfed families. The broad-sense heritability accounted for 33–43% of the variation in PA levels. Families also differed significantly in the amount and the direction of PA induction in both shoots and roots, 24 h after punching 15 holes in the leaves. We found a significantly negative relationship between the changes in PA content of the shoots and changes in PA content of the roots. The total PA content of the plants did not increase. We thus concluded that changes in PA distribution over the plant resulted from transport of PAs within the plant. The direction of transport differed between families: some transported PAs to the shoots, others to the roots. This makes it questionable whether PAs act as damage-induced defences. The effect of damage on the PA concentration is far less than the differences found between families in the constitutive PA concentration. This again strongly suggests that damage-induced defences inCynoglossum officinale do not play an important role. We argue that the general lack of attention that is given to genotype in induction experiments, has led to false conclusions.     

Leaf development of the ostrich fern Matteuccia struthiopteris (L.) Todaro

The timing of emergence of the three different leaf types of Matteuccia struthiopteris is described from plants sampled over the course of a growing season. Vegetative leaves were first to appear, followed five weeks later by sporophylls and cataphylls. Leaf number and type, and total leaf dry weight per plant were assessed in weekly transects. Vegetative fronds contributed the most to total leaf dry weight, which increased during the first four weeks, and then remained constant for the remainder of the season. Cataphylls, although numerous by the end of the season, contributed little weight. Sporophylls occurred on the widest plants with the most vegetative leaves and greatest leaf weight, whereas cataphylls occurred on most plants except the smallest. Experimentally defoliated plants were re-examined in late summer. Following initial harvest, plants often produced a second smaller set of leaves. These were restricted to vegetative leaves and cataphylls. Ability to reissue leaves, especially vegetative fronds, declined very quickly after the first few weeks in the growing season. Defoliated plants draw on the extensive reservoir of developing leaves which are found on the rhizome, thus possibly diminishing the ability of the plant to withstand regular harvesting of the young fronds for food. Individual leaves were tagged and measured over the growing season. Non-linear regression curves fitted to the growth data for the three types of leaves indicate that growth was described best by a monomolecular growth curve for the vegetative and fertile fronds. Cataphyllar growth could be described equally well by either a monomolecular or a logistic function.     

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.     

Phenology and water relations of tree sprouts and seedlings in a tropical deciduous forest of South India

The phenology of sprouts (>1 year old, up to 1.5 m in height) and seedlings (<1 year old) of six woody species (four deciduous, one brevi-deciduous, and one evergreen) was examined during the dry season in a tropical deciduous forest of South India. Xylem water potential (_x), leaf relative water content (RWC; % turgid weight), and xylem specific conductivity (K _S; kg s^–1 m^–1 MPa^–1) of sprouts were measured on two occasions during the dry season. In addition, K _S of seedlings (<1 year old) of one deciduous and one evergreen species was determined to allow comparison with sprouts. _x of deciduous species was significantly higher at the second sampling date and was accompanied by a significant increase in K _S and RWC, while the brevi-deciduous and evergreen species did not show any difference in _x. Seedlings of Terminalia crenulata (deciduous) and Ixora parviflora (evergreen) had significantly lower K _S compared to sprouts, while seedlings of all four deciduous species shed their leaves much earlier in the dry season than did conspecific sprouts. More favorable water relations of sprouts compared to seedlings during the peak of the dry season may explain the lower rates of die-back and mortality of sprouts observed in dry deciduous forests of India.

---

This revised version was published online in May 2005 with corrections to Received-/Accepted-dates.     

Polyamine levels in buds and twigs of Tilia cordata from dormancy onset to bud break

The fluctuations of free and bound polyamines (PAs) were studied in vegetative buds and underlying twigs of linden (Tilia cordata L.) from August to May, to assess the connection between PA levels and seasonal cycles of growth and dormancy. Outer and inner bud scales and shoot tips (short shoot tips with leaf initials in contiguous short internodes) were analyzed separately, as were phloem with cortex and xylem with pith tissue from twigs. Seasonal variations in PA levels were present in buds and twigs during the research period. The most abundant PA in buds and twigs in free and bound forms was spermidine followed by putrescine. PA amounts were low in buds and twigs in autumn. In twig tissues, free PAs were predominant whereas in bud scales, bound PAs accumulated over free PAs in autumn, first in inner scales and later in outer scales as well. PA levels did not increase dramatically during the onset of dormancy in autumn but lower temperatures and probable cold hardening correlated positively with bound PAs in bud scales. In shoot tips with leaf initials, and contiguous short internodes, free putrescine and spermidine levels rose simultaneously with bud burst and new growth, while bound PAs diminished quite radically from temporary bud scales and from growing shoot tips.     

Seasonal ectomycorrhizal fungal biomass development on loblolly pine (Pinus taeda L.) seedlings

Ergosterol, a membrane sterol found in fungi but not in plants, was used to estimate live mycelial biomass in ectomycorrhizae. Loblolly pine (Pinus taeda L.) seeds were sown in April 1993 and grown with standard nursery culture practices. Correlations between total seedling ergosterol and visual assessment of mycorrhizal colonization were high during July and August but low as ectomycorrhizal development continued into the growing season. Percentages of mycelial dry weight over lateral roots decreased from 9% in July to 2.5% in November because seedling lateral root dry weight accumulated faster than mycelial dry weight. Total ergosterol per seedling increased from July through February. As lateral root dry weight ceased to increase during winter months, ectomycorrhizal mycelia became the major carbohydrate sink of pine seedlings. No distinctive seasonal pattern of soil ergosterol content was observed. The impact of ectomycorrhizal fungi on plant carbohydrate source-sink dynamics can be quantitatively estimated with ergosterol analysis but not with conventional visual determination.     

Seasonal variation in honeyeater foraging behaviour,infloresence abundance and fruit set in Banksia spinulosa (Proteaceae)

Seasonal variation in the foraging behaviour of honeyeaters and the production of fruit were examined in relation to the flowering intensity of Banksia spinulosa over two flowering seasons. The abundance of inflorescences was greater in the mid than in the early and late periods of the flowering season. In the mid period, many plants were blooming and each plant had many flowering inflorescences. Inflorescences received most visits by honeyeaters in the early flowering period; the visitation rate declining as flowering progressed. Eastern spinebills were the most common floral visitors at all times during the season. The number of foraging probes made at inflorescences by eastern spinebills did not differ throughout the season. Foraging movements between inflorescences on the same plant were more frequent in the mid period than in the early and late periods. Long distance movements between plants (more than 10 m apart) were promoted by aggressive interactions between honeyeaters. Inflorescences flowering in the late period were less likely to develop follicles because there were fewer visits by birds and/or because resources had been allocated to inflorescences pollinated earlier in the season. The number of follicles produced per infructescence did not differ between flowering periods. Overall, the number of inflorescences produced per plant, the number of visits received per inflorescence and the proportion of inflorescences that developed follicles were greater in 1987 than in 1988.     

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     

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.     

The seasonal pattern of CO2 exchange of Festuca rubra L. in a montane meadow community in Northern Germany

Summary Completely climatized cuvettes were used to follow the CO₂ gas exchange of red fescue (Festuca rubra L.), growing on a fertilized and an unfertilized plot, during a growing season from May through October. Objective of the study was to determine the effect of environmental factors on the seasonal CO₂ gas exchange.Gas exchange rates were calculated on the basis of leaf dry weight, surface area and chlorophyll content. Photosynthetic rates differed between the fertilized and unfertilized plants when based on leaf dry weight or leaf surface area but were similar when based on chlorophyll.Multiple regression analysis was used to related photosynthetic rates to radiation, temperature, water vapor concentration difference, chlorophyll content and time. A cubic regression equation based on daily radiation alone explained 85% of the variation for the fertilized plants and 87% of the variation for the unfertilized plants.During the growing season the unfertilized plants had a continual decline in their photosynthetic rates. The fertilized plants had high photosynthetic rates in the spring and in the fall.Light response curves indicated greater photosynthetic rates at light saturation as well as in the light limited portion of the light response curve for the fertilized plants. Photosynthetic rates of the fertilized plants were generally depressed during periods of warm temperature and high light intensity in June and July.Photosynthetic rates declined at temperatures above 24°C. The decline was greater for the more mesomorphic fertilized plants. A similar response was noted to increasing water vapor difference, although it was difficult to separate from the temperature effect. Maximum photosynthetic rates were found between 14°C and 22°C, although there was considerable variation in the maximum rates.The effects of cutting (mowing) on the gas exchange were difficult to determine due to the interaction of the environmental factors.Chlorophyll content showed significant correlation with photosynthetic rates.     

Spermidine and related-metabolic inhibitors modulate sugar and amino acid levels in Vitis vinifera L.: possible relationships with initial fruitlet abscission

The relationships between free polyamines (PAs), sugar and amino acid status were investigated in cuttings from two cultivars of Vitis vinifera L., Pinot noir (PN), a low abscising cultivar and Merlot (MRT), a high abscising one. In both cultivars free PAs decreased in inflorescences, but more drastically in MRT plants. Upon anthesis, this was associated with a decreased sugar content, especially sucrose, and an increase in total free amino acids. Thereafter, sucrose and amino acids showed opposite trends. In addition, darkening the PN plants at full flowering resulted in a dramatic decrease of PAs and sugars in inflorescences, but an increase in amino acid content, followed by high abscission. The concept that initial fruitlet abscission might be regulated by free PAs through changes in primary metabolites was hypothesized. Hence, the application of exogenous spermidine (Spd), but not putrescine (Put), prior to flowering markedly inhibits abscission. The Spd treatment also increased soluble sugar content but reduced amino acids in both leaves and inflorescences, while Put had no significant effect. By contrast, cyclohexylamine and beta-hydroxyethylhydrazine, as potent inhibitors of Spd synthase and PA-oxidases, respectively, exerted inverse effects on sugar, amino acid and abscission levels. Sucrose and free proline seemed to be highly sensitive to these treatments. This study suggests that Spd could regulate fruitlet abscission in grapevine by modulating, in a reverse way, the levels of sugars and amino acids in inflorescences.     

Studies on the Growth, Flowering, and Production of Female Sterile Flowers as Affected by Different Levels of Foliar Potassium in Solanum sisymbrifolium Lam.: I. EFFECT OF POTASSIUM CONTENT OF THE PLANT ON VEGETATIVE GROWTH AND FLOWERING

As compared to a high level of foliar potassium, a low but nota deficient level promotes shoot elongation and flowering inS.sisymbrifolium.This is accompanied by formation of flowers with rudimentaryovaries in which the megagametophyte in the ovules aborts atthe two- or the four-nucleate stage. Plants having a higherK content do not bear the female sterile flowers. Inflorescencesof the high-K plants with 20 per cent fewer buds have 23 percent higher dry weight than inflorescences of low-K plants.The ratio of organic nitrogen to dry weight is not affected,indicating that the inflorescences of the high-K plants arebetter supplied with metabolites in general.     

Occurrence and biological significance of proanthocyanidins in the American diet

Dietary intake of proanthocyanidins (PAs) has been largely unknown because of the lack of reliable values for their content in foods. Recent development of an analytical method for PAs has allowed the quantification of individual oligomers and polymers. This method has been employed to analyze food samples collected under the USDA National Food and Nutrition Analysis Program. A database of the PA content in common foods and also infant foods has been established. It has been shown that PAs account for a major fraction of flavonoids ingested in the US diet and infants and children appear to ingest more PAs than adults on the basis of body weight. These data will provide an opportunity to examine the association between PA intake and health and disease outcomes in epidemiological studies. PA analysis and the significance of PAs in nutrition and diet are reviewed.     

Effect of pollen limitation, additional nutrients, flower position and flowering phenology on fruit and seed production in Salvia verbenaca (Lamiaceae)

Variation in fruit set, seed/ovule ratio and mean single-seed weight among plants, and within plants among inflorescences, was investigated in a population of Salvia verbenaca in northwest Spain. Mean single-seed weight varied significantly among plants which flowered at about the same time and mean single-seed weight was significantly greater for plants that flowered late in the season than for plants flowering early in the season. Within individual plants, mean single-seed weight increased steadily between the earlier opening basal inflorescences and the later opening upper inflorescences. However, neither fruit set nor seed/ovule ratio varied significantly among inflorescences. Previous work on this population indicates that outcrossing improves mean single-seed weight, so I suggest that the observed pattern of among-inflorescence variation in mean single-seed weight may be attributable to a gradual increase in nonself pollen receipt as the flowering season progresses.     

Spatial and temporal distribution of the alkaloid sanguinarine in Sanguinaria canadensis L. (Bloodroot)

Acclimated and non-acclimated potted plants of Sanguinaria canadensis L. were harvested at early and late dormancy, anthesis, and immature and mature fruiting stages. Sanguinarine content and concentration were determined for rhizomes (distal, proximal, and middle sections), roots, leaves, flower, and fruit. Rhizomes had highest sanguinarine content and concentrations, and exhibited decreasing concentration gradients from the distal to proximal third. Concentrations in roots were a tenth of rhizome concentration. Concentrations in leaves, flowers, and fruit were one-thousandth of rhizome Sanguinarine content in whole acclimated plants was constant. Content in whole nonacclimated plants increased as the plant became physiologically active, but was constant during fruit maturation: content in roots, leaves, and fruit did not change. The substantial increase in whole-plant dry weight coupled with the unchanging sanguinarine content during fruit maturation suggests either a shift in photosynthate allocation from defense to growth, or a constant turnover of sanguinarine.     

Lead Content in Pot Marigold (<Emphasis Type="Italic">Calendula officinalis</Emphasis> L.) Inflorescences and Leaves: Impact of Precipitations and Vicinity of Motorway

Trace metal contamination is a major environmental and health problem virtually in all countries. The present study was aimed to estimate the lead content of pot marigold (Calendula officinalis L.) inflorescences and leaves collected from a nonpolluted test field. The lead content in dry pot marigold inflorescences was 9.34 ± 0.79 μg/g, in dry leaves 11.57 ± 0.47 μg/g, and in soil 0.649 ± 0.012 μg/g. The distance of pot marigold collection beds (30–220 m from the motorway) had no effect on lead content. There was a strong positive correlation between the amount of precipitations and lead content of pot marigold leaves but not inflorescences indicating the soil as primarily the source of increased lead content. In conclusion, no effect of motorway vicinity was found for pot marigold inflorescences or leaves lead content; however, as a precaution, it is not recommended to collect the plants during or just after showers.     

Leaf development of the ostrich fern Matteuccia struthiopteris (L.) Todaro

The timing of emergence of the three different leaf types of Matteuccia struthiopteris is described from plants sampled over the course of a growing season. Vegetative leaves were first to appear, followed five weeks later by sporophylls and cataphylls. Leaf number and type, and total leaf dry weight per plant were assessed in weekly transects. Vegetative fronds contributed the most to total leaf dry weight, which increased during the first four weeks, and then remained constant for the remainder of the season. Cataphylls, although numerous by the end of the season, contributed little weight. Sporophylls occurred on the widest plants with the most vegetative leaves and greatest leaf weight, whereas cataphylls occurred on most plants except the smallest. Experimentally defoliated plants were re-examined in late summer. Following initial harvest, plants often produced a second smaller set of leaves. These were restricted to vegetative leaves and cataphylls. Ability to reissue leaves, especially vegetative fronds, declined very quickly after the first few weeks in the growing season. Defoliated plants draw on the extensive reservoir of developing leaves which are found on the rhizome, thus possibly diminishing the ability of the plant to withstand regular harvesting of the young fronds for food. Individual leaves were tagged and measured over the growing season. Non-linear regression curves fitted to the growth data for the three types of leaves indicate that growth was described best by a monomolecular growth curve for the vegetative and fertile fronds. Cataphyllar growth could be described equally well by either a monomolecular or a logistic function.     

Tissue distribution, core biosynthesis and diversification of pyrrolizidine alkaloids of the lycopsamine type in three Boraginaceae species

Three species of the Boraginaceae were studied: greenhouse-grown plants of Heliotropium indicum and Agrobacterium rhizogenes transformed roots cultures (hairy roots) of Cynoglossum officinale and Symphytum officinale. The species-specific pyrrolizidine alkaloid (PA) profiles of the three systems were established by GC-MS. All PAs are genuinely present as N-oxides. In H. indicum the tissue-specific PA distribution revealed the presence of PAs in all tissues with the highest levels in the inflorescences which in a flowering plant may account for more than 70% of total plant alkaloid. The sites of PA biosynthesis vary among species. In H. indicum PAs are synthesized in the shoot but not roots whereas they are only made in shoots for C. officinale and in roots of S. officinale. Classical tracer studies with radioactively labelled precursor amines (e.g., putrescine, spermidine and homospermidine) and various necine bases (trachelanthamidine, supinidine, retronecine, heliotridine) and potential ester alkaloid intermediates (e.g., trachelanthamine, supinine) were performed to evaluate the biosynthetic sequences. It was relevant to perform these comparative studies since the key enzyme of the core pathway, homospermidine synthase, evolved independently in the Boraginaceae and, for instance, in the Asteraceae [Reimann, A., Nurhayati, N., Backenkohler, A., Ober, D., 2004. Repeated evolution of the pyrrolizidine alkaloid-mediated defense system in separate angiosperm lineages. Plant Cell 16, 2772-2784.]. These studies showed that the core pathway for the formation of trachelanthamidine from putrescine and spermidine via homospermidine is common to the pathway in Senecio ssp. (Asteraceae). In both pathways homospermidine is further processed by a beta-hydroxyethylhydrazine sensitive diamine oxidase. Further steps of PA biosynthesis starting with trachelanthamidine as common precursor occur in two successive stages. Firstly, the necine bases are structurally modified and either before or after this modification are converted into their O(9)-esters by esterification with one of the stereoisomers of 2,3-dihydroxy-2-isopropylbutyric acid, the unique necic acid of PAs of the lycopsamine type. Secondly, the necine O(9)-esters may be further diversified by O(7)- and/or O(3')-acylation.