Primula Veris Research Articles

Near-infrared diffuse reflection spectroscopy and multivariate calibration hyphenated with thin-layer chromatography for quality control of a phytomedicine and simultaneous quantification of methoxylated flavones

By use of near infrared (NIR) spectroscopy hyphenated to thinlayer chromatography (TLC) for analysis of methoxylated flavones in a phytomedicine we sought to achieve two objectives: first, to establish a method for rapid, qualitative identification of five methoxylated flavones, denoted G1, G2, G3, G4, and G5, in order of their RF values in normal-phase TLC, and, second, to produce a quantitative model for analysis of G4 (3′,4′,5′-trimethoxyflavone), the compound most representative of Primula veris flowers in phytomedicine. To provide appropriate reference analytical data for building the multivariate cluster and partial least-squares regression (PLS) model, TLC was performed on alumina with n-hexane-ethyl acetate 70:30 (v/v) as mobile phase. Forty-four spectra of eleven independent phytomedicine samples were analyzed with five scans to generate a qualitative cluster model based on PCA (principle-components analysis) that enabled differentiation between G1-G5 on the basis of their methoxylation pattern. ...

Simulating direct and indirect effects of climatic changes on rare perennial plant species in fragmented landscapes

Question: How does climate change influence plant species population dynamics, their time to extinction, and proportion of occupied habitats in a fragmented landscape? Location: Germany and Central European lowland. Methods: We apply a mechanistic general simulation model to test the response of plant functional types to direct and indirect effects of climate change. Three functional types were chosen to represent a set of well-studied perennial plant species: Juncus atratus, Gentiana pneumonanthe and Primula veris. We link local population dynamics within a heterogeneous, fragmented landscape context. “Species spheres”, i.e. multi-dimensional parameter ranges rather than single parameter realizations, based on field and literature data served as proxy for life stage transition parameters. Four climatic scenarios summarizing different cumulative weather effects on demographic rates and different local disturbance frequencies were run. The model predicts “time to extinction” (TE) and “proportion of occupied habitat” (POH) as regional indicators for species extinction risk. Results: TE decreased for all species when weather conditions worsened, and even more so when the frequency of local destructive events additionally increased. However, management towards fewer disturbance events could buffer the negative effect of climate to some extent. The magnitude of these responses varied with species type. POH declined with an increase in bad weather as well as with increasing disturbance frequency. The better the climatic conditions, the less severe were disturbances on population performance. Conclusions: The “species spheres” proved to be a valuable approach for predictive trends. As climate change usually also implies destructive events such as land-use change, flooding or fire, our model on local and regional extinction risks can support conservation issues and management actions.

Integrated treatment of cardiovascular common pathologies with conventional pharmacological therapy integrated with a typical remedy of anthroposophic medicine, Cardiodoron (Onopordon/Primula comp)

Anthroposophic medicine (AM) in Italy is mostly practiced in private settings: practices, multidisciplinary day-centers and a thermal clinic. In the General Medicine Department of the Istituto Clinico San Siro (ICSS), in Milan, AM has been used since 2000 in an integrated context, as a complementary or alternative treatment to conventional therapies. The department has one ambulance and 10 beds and treats acute, subacute and chronic orthopedic, postoperative, cardiovascular and oncological pathologies. Approximately 250 patients are treated each year. ICSS is a private clinic operating under a convention with the Italian NHS. The department medical team practices also anthroposophic medicine. At the beginning of XX century, the typical remedy Cardiodoron (a special composition of Onopordon acanthium, Primula officinalis, Hyoscyamus niger) was introduced in the context of AM for regulating cardiovascular system. Typical remedies have been conceived as image and support of the healthy balance of an organ, or an organ system or of a human complex function. In the ICSS department, patients with diagnose of heart failure, arrhythmia and hypertension, after giving their consent, receive systematically also oral Cardiodoron as a long-term regulating treatment. Since 2000, over 2000 patients have been treated and registered in the department. Both conventional and anthroposophical treatments are paid by the Italian NHS. The compliance to AM is high. To have an assessment of Cardiodoron use, 30 records of inpatients with heart failure (NYHA II/III) from period 2002 to 2003 have been randomly selected and statistically analysed in a pilot epidemiological–clinical observational study. The 30 patients (19 female, 11 male), mean age 78.9–78.5, respectively, were monitored with instant and Holter ECG, arterial pressure and cenesthesis. Improvement of these parameters and a quick disappearance of fear and anxiety were reported in all patients. In the present study, it was not possible to differentiate the effect of conventional and anthroposophical treatments, administered together. The hypothesis is of an optimising therapeutic effect of conventional therapies given by Cardiodoron. The department records since 2000 are being included in a database in order to have an overview of the value added of Cardiodoron treatment. Preliminary results will be presented.

Antiviral activity of a composition of Gentiana lutea L., Primula veris L., Sambucus nigra L., Rumex spec. andVerbena officinalis L. (Sinupret®) against viruses causing respiratory infections

Sinupret ® , an aqueous-ethanolic extract from five plants, Gentiana lutea L., Primula veris L., Sambucus nigra L., Rumex spec. and Verbena officinalis L., is frequently used in the treatment of acute and chronic rhinosinusitis and respiratory viral infections such as common cold. To date little is known about basic mechanisms of its potential antiviral activity. Therefore experiments have been performed for the detection of its antiviral activity against a broad panel of human pathogenic enveloped and non-enveloped RNA and DNA viruses causing infections of the upper respiratory tract: influenza A virus, parainfluenza virus, human rhinovirus B, coxsackievirus, adenovirus C, and respiratory syncytial virus. Determination of virus production was performed after treatment of the infected cells with non-toxic concentrations of the commercially available product using plaque-reduction assays, analyses of cytopathogenic effects and ELISAs for viral proteins. Antiviral activity of Sinupret ® could be detected independent of the type of the viruses in RNA as well as in DNA virus infected cell cultures and also against coated and uncoated viruses. A very strong inhibitory activity was obvious against adenovirus and respiratory syncytial virus infections. These results demonstrate that Sinupret ® showed a broad antiviral activity which basic mechanisms are still unclear. This has to be elucidated in further studies.

Hydrocarbon Footprints as a Record of Bumblebee Flower Visitation

Bumblebees leave traces of cuticular hydrocarbons on flowers they visit, with the amount deposited being positively related to the number of visits. We asked whether such footprint hydrocarbons are retained on flowers for sufficiently long periods of time so as to reflect bee visitation in pollination studies. In laboratory experiments, flower corollae (Primula veris, Digitalis grandiflora) visited by Bombus terrestris workers retained bee-derived nonacosenes (C(29)H(58)) in near-unchanged quantities for 24 hours, both at 15 and 25 degrees C. Additionally, synthetic (Z)-9-tricosene applied to flower corollae of the deadnettle Lamium maculatum was retained for 48 hours in an unchanged quantity. In a field survey, the amount of footprint alkenes on flowers of comfrey (Symphytum officinale) plants was positively correlated with the number of bumblebee visits that those plants had received during the day. Together, these data suggest that flowers retain a long-term quantitative record of bumblebee visitation. The analysis of petal extracts by gas chromatography could provide a cheap and reliable way of quantifying bumblebee visits in landscape scale studies of pollination.

Environmental context drives seed predator-mediated selection on a floral display trait

Linking trait selection to environmental context is necessary to move beyond the simple recognition that selection is spatially variable and to understand what ultimately drives this variation. Natural selection acts through differences among individuals in lifetime fitness and information about effects on fitness components is therefore often not sufficient to gain such an understanding. We investigated how environmental context influenced intensity of seed predation, flower abortion and selection on floral display traits in 44-52 populations of the perennial herb Primula veris over 2 years. Phenotypic selection on both inflorescence height and flower number varied among populations and was mediated partly by pre-dispersal seed predation and flower abortion in one of the years. Among-population variation in selection on inflorescence height, but not flower number, was linked to variation in canopy cover via its effects on seed predation. Lifetime fitness was less sensitive to seed predator damage in shaded environments but estimates of selection based on lifetime fitness agreed qualitatively with those based on seed output. Our results demonstrate that seed predators constitute an important link between environmental conditions and trait evolution in plants, and that selection on plant traits by seed predators can depend on environmental context.

The contents of therapeutically effective compounds of cowslip (Primula veris L.) from various stands of Levocske Mountains in eastern Slovakia

The contents of therapeutically effective compounds of cowslip (Primula veris L.) from various stands of Levocske Mountains in eastern Slovakia

Biological Flora of the British Isles: Primula veris L.

Summary This account presents information on all aspects of the biology of Primula veris that are relevant to understanding its ecological characteristics and behaviour. The main topics are presented within the standard framework of the Biological Flora of the British Isles: distribution, habitat, communities, responses to biotic factors, responses to environment, structure and physiology, phenology, floral and seed characters, herbivores and diseases, history and conservation. Primula veris is a distylous, perennial rosette hemicryptophyte, mostly found in well‐drained, herb‐rich meadows and grasslands, in scrub or woodland rides and edges, and on calcareous cliffs. It occurs on base‐rich loams or clays, on limestone, and sometimes in dunes. Native to the British Isles, it is characteristic of many mesotrophic and calcicolous lowland grassland communities, but it also extends into upland valleys in northern England and Scotland. In Ireland it is also native and most abundant in the central plain. Primula veris occurs throughout the temperate areas of Europe as far east as the Russian border. Primula veris is a shade‐intolerant species that shows reduced performance and flowering under shade or increased competition. The species is not able to penetrate anoxic soil layers and is therefore mostly absent from locations that are characterized by a high water table. It has a well‐developed drought tolerance. Primula veris is an obligate outbreeder which entirely depends on foraging insects for successful pollination. Although the species shows heterostyly, with two reciprocal morphs (pin and thrum), only relatively low levels of intermorph pollination have been reported. Seed dispersal is restricted to a few centimetres from maternal plants, whereas pollen flow is wider, but still limited to a few metres from parental plants. Both are factors that contribute to a significantly fine‐scale spatial genetic structure and small neighbourhood size. At several locations throughout the British Isles, P. veris occurs together with P. elatior and especially P. vulgaris. The hybrid P. veris × P. vulgaris = P. × polyantha occurs frequently in mixed populations, whereas the hybrid P. veris × P. elatior = P. × media has been rarely reported in Britain. Although P. veris is still a widespread grassland herb, it is less abundant in the British Isles and continental Europe than before. This decline can be attributed to changes in land‐use practices, such as the loss of traditional hay–meadow management, the loss of grazing and an increase of ploughing, in combination with ongoing destruction and nutrient enrichment of permanent grasslands. However, from 1980 onwards the species has increased substantially in many areas of the UK, largely because its seeds are included in wildflower seed‐mixtures that are sown on new or upgraded road and motorway verges, embankments and urban conservation areas.

Antimitotic and antibacterial effects of the Primula veris L. flower extracts

Abstract Primula is a plant genus which comprises about 400 species. It has been found in a number of pharmacological studies that primrose extracts are rich in saponins. Phenolic glycosides and saponins are characteristic compounds for the genus Primula. In this study several flower extracts from Primula veris L. has been tested for antibacterial activity and decoction from the flowers has been tested for antimitotic activity. Antibacterial activity was determined by the well diffusion method and Allium cepa L. has been used for evaluating cytotoxicity. Decoction of flowers was toxic on root number and root length in A. cepa L. and reduced the mitotic index significantly. All of the tested P. veris L. extracts showed inhibitory effect against both Gram positive and Gram negative microorganisms at varying degrees. The most effective fraction was found to be the ethanolic.

Saponins in tissue culture of Primula veris L.

Roots of Primula veris L. contain considerable amounts of triterpene saponins, which are used in medicine as expectorants. P. veris is in many places an endangered plant, and its production in the field is laborious and a low yielding process. Plant tissue culture provides an alternative means for producing secondary metabolites. Shoot apex, callus, suspension, and root cultures of P. veris were developed for saponin production. In these cultures, the content of triterpene saponins, with focus on primula acid I, the most dominant saponin in Primula species, was determined and compared to that in soil-grown plants. The highest content of primula acid I was observed in root cultures, on average 29.5 mg/g dry weight. Some culture lines contained higher amounts of primula acid I (62.6 mg/g dry weight) than the roots of plants grown in soil.

Morph-specific variation of floral traits associated with reciprocal herkogamy in natural populations of Primula vulgaris and Primula veris

Documenting the morph-specific variation of floral traits associated with reciprocal herkogamy is of special importance for revealing the functional significance of traits in the evolution and maintenance of the heterostylous syndrome. In order to describe the extent and specificity of variation, stigma height, anther height, stigma-anther separation and corolla tube length were measured on 800 flowers collected in two natural populations of Primula vulgaris and P. veris. Beside the almost complete separation of stigma heights between the two morphs, we found appreciable intermorph overlap in anther height and relatively broad range of stigma-anther separation so far reported for heterostylous species. The corolla tube length–stigma-anther separation relationship showed striking difference between the two floral morphs, which supports the hypothesis that length of the corolla tube plays a more important role in positioning the sex-organs in the appropriate distance in the short-styled than in the long-styled morph.

Fine‐Scale Spatial Genetic Structure of the Distylous Primula veris in Fragmented Habitats

In Flanders (northern Belgium), the distylous self-incompatible perennial herb Primula veris is common, but mainly occurs in fragmented habitats. Distyly, which favours disassortative mating, is characterized in P. veris by two genetically determined floral morph types (pin or thrum). Using 18 polymorphic loci, we investigated fine-scale spatial genetic structure (SGS) and spatial distribution of the morphs within four populations from two regions that differ in degree of habitat fragmentation. We studied the contributions made by sexual reproduction and clonal propagation and compared the SGS patterns between pin and thrum morph types. Clonal growth was very restricted to a few individuals and to short distances. One population showed a non-random spatial distribution of the morphs. Pin and thrum individuals differed in SGS patterns at a small scale, suggesting intrapin biparental inbreeding, also related to high plant densities. This may be explained by partial self-compatibility of the pin morph combined with restricted seed dispersal and pollinator behaviour. There is an indication of more pronounced SGS when populations occur in highly fragmented habitats. From our findings, we may hypothesize disruption of the gene flow processes if these large populations evolve into patchworks of small remnants, but also a possible risk for long-term population survival if higher intrapin biparental inbreeding leads to inbreeding depression. Our study emphasizes the need for investigating the interactions between the heterostylous breeding system, population demographic and genetic structure for understanding population dynamics in fragmented habitats and for developing sustainable conservation strategies.

Environmental context influences the outcome of a plant–seed predator interaction

Mutualistic and antagonistic interactions with animals are known to influence the performance of plants in many ways. Much less is known about how such effects are influenced by the environment and how they translate into effects on plant population dynamics. In this study, we first quantified how pre‐dispersal seed predation in the perennial herb Primula veris changes along a continuous gradient of canopy cover. We then used a deterministic demographic modeling approach to investigate how seed predation may influence population growth rate (λ) and how effects depend on environmentally‐induced variation in plant demography. Intensity of seed predation increased with the degree of canopy cover, while sensitivity of λ to changes in seed production decreased. This translated into non‐linear effects of seed predation on λ along the canopy cover gradient. Despite seed predation rates being highest in closed habitats, the negative effect of seed predation on λ was lower here than in slightly more open habitats. Our results demonstrate that knowledge of the intensity of plant‐herbivore interactions does not suffice to infer how animals influence the population dynamics of plants and their distribution. Plant demographic sensitivity and its dependence on the environment need also to be taken into account when assessing the importance of plant–animal interactions.

Morph-specific differences in reproductive success in the distylous Primula veris in a context of habitat fragmentation

Abstract Heterostylous self-incompatible plant species are particularly sensitive to habitat fragmentation and to disruption of pollination processes because of the need of intermorph cross-pollination for producing seeds. Heterostyly is characterized by sexual polymorphism through the occurrence of two (distyly) or three (tristyly) morph types that differ in floral traits (style length and anther position). We examined whether the long-styled (pin) and short-styled (thrum) morph types show differences in reproductive components and responses to habitat fragmentation in the distylous, self-incompatible perennial herb Primula veris. We documented reproductive components for pin and thrum individuals and their relationships with population size, plant density and morph ratio (pin frequency), in nine populations from Flanders (northern Belgium) located in fragmented habitats of the intensively used agricultural landscape. Seed abortion increased in small populations as a result of inbreeding depression. Fruit set increased with plant density. Seed set was positively related to pin proportion. Seed set was higher for pin than thrum in small populations, but lower in large populations. Two hypotheses can be considered to explain these morph-specific differences: a pollen transfer asymmetry, and a reproductive advantage for the partially self-compatible pin morph. Morph types appear to respond differently to habitat fragmentation constraints. A floral morph type showing partial self-compatibility may be favored in populations under pollination failure, because it can increase reproductive success and mating opportunities through intramorph crosses.

Is the colour dimorphism inDactylorhiza sambucina maintained by differential seed viability instead of frequency-dependent selection?

The European rewardless, bee-pollinated orchid Dactylorhiza sambucina commonly produces yellow-flowered and purple-flowered individuals in frequencies that range from balanced (per population) to very unbalanced, with parts of the species' range entirely monochromatic. We studied male and female reproductive success of the two morphs in 22 populations in the Czech Republic, relating it to morph frequency, population size and density, and presence and abundance of yellow and purple co-flowering nectar-providing species visited by the same bee species. Cumulative abundances of yellow nectar-producing co-flowering species (of which, on average, Primula veris made up 56%) had a negative effect on male reproductive success of the yellow morph, and spectral analyses showed that to bumblebees the colours of P. veris and yellow D. sambucina are different, permitting ready visual discrimination. The cumulative abundance of purple co-flowering species had no significant effect on morph reproductive success. Morph frequencies were unrelated to reproductive success and population size, and there was no evidence of frequency-dependent selection except in one highly unbalanced population. Density of flowering conspecifics was negatively correlated with male reproductive success of the purple morph. Seed mass, viability, and germination success depended on whether seeds resulted from outcrossed or selfed matings and on morph colour. Selfed seeds and seeds produced by the yellow morph from yellow yellow and yellow purple crosses had zero germination (after three months), providing the first hint that differential vegetative fitness, rather than differential reproductive fitness via pollinator selection, may explain morph frequencies in D. sambucina.

Habitat Change and Demography of Primula veris: Identification of Management Targets

Although the effects of deterministic factors on population viability often are more important than stochasticity, few researchers have dealt with the effect of deterministic habitat changes on plant population demography. We assessed population viability for the perennial herb Primula veris L. and identified targets for management based on demographic data from five different habitat types representing different degrees of canopy closure. We conducted replicate studies at the border of the distribution area and in more central parts. Demographic patterns were similar between the two regions. Most study populations had a positive population growth, and only populations in late phases of forest succession showed consistently negative trends. The populations of open habitats had high seedling recruitment, and the populations of early and middle forest succession had high seed production. The importance of survival for population growth rate increased with increasing habitat closure, whereas the importance of growth and reproduction decreased. Results of the elasticity analysis suggested that the best method to manage decreasing late-successional populations is to increase survival of the largest individuals. The life-table response experiment (LTRE) analysis, however showed that survival of the largest individuals contributed little to differences in population growth rates of different habitats, whereas seed production and growth of small individuals were more important. Moreover direct perturbation of the performance of the largest stages showed that late-successional populations would not attain positive population growth even if the largest stages had no mortality at all. We conclude that restoration of recruitment is the only possibility for positive population growth in late-successional populations of P. veris, although the elasticities of recruitment transitions are low. Our results also suggest that retrospective demographic methods such as LTRE constitute an important and necessary complement to prospective methods such as elasticities in identifying management targets.

Analysis of phenolic glycosides and saponins in Primulaelatior and Primulaveris (primula root) by liquid chromatography, evaporative light scattering detection and mass spectrometry

This paper describes the first liquid chromatographic method suitable for the simultaneous determination of bioactive compounds, saponins and phenolic glycosides, present in Primula elatior and Primula veris, including the NMR data of primulaverin and primeverin. Optimum separations were obtained with a Synergi 4 μm Fusion RP 80 Å column, using 0.025% TFA in water and 5% acetonitrile in methanol as mobile phase. Saponins were detected by evaporative light scattering detection (ELSD), whereas the phenolic glycosides were monitored by UV at 210 nm. The method was validated for repeatability ( σ rel ≤ 4.5%), precision (intra- and inter-day variation ≤5.0%), accuracy (recovery ≥97.1%) and sensitivity (LOD ≤22 ng (UV) and ≤38 ng (ELSD) on-column, respectively). LC–MS experiments in negative APCI mode allowed a final peak assignment. Both Primula species could easily be differentiated by their saponin pattern. The total saponin content was highest in P. veris roots (max. 14.9%), the aerial parts or P. elatior contained significantly less amounts; primeverin (0.64–1.42%) showed to be the most dominant phenolic glycoside.

Screening of plants used in Danish folk medicine to treat epilepsy and convulsions

Aqueous and ethanolic extracts of 42 plants used in Danish folk medicine for the treatment of epilepsy and convulsions, or for inducing sedation, were tested for affinity to the GABA A–benzodiazepine receptor in the flumazenil-binding assay. Ethanolic extracts of leaves of Primula elatior and Primula veris and aerial parts of Tanacetum parthenium exhibited good, dose-dependent affinity.

Within-population genetic variation in the distylous Primula veris: Does floral morph anisoplethy matter in fragmented habitats?

We studied genetic variation and structure in 24 populations of Primula veris , a distylous self-incompatible perennial herb that in Flanders (northern Belgium) occurs in fragmented habitats of the intensively used agricultural landscape. Distyly, which favours cross-fertilization, is characterized in this species by two genetically determined floral morph types (‘pin’ or ‘thrum’). Using 30 allozyme loci, we investigated within-population and within-morph genetic variation and structure in relation to distyly (stochastic morph bias and pin frequency), demography (population size, plant density), and evidence of recent bottlenecks. No significant relationship was found between genetic patterns and stochastic morph bias. However, F IS values were lower when pin frequency increases in the population. This may be explained by the partial self-compatibility of the pin morph, which, in conditions of pollination disruption, as observed for P. veris in fragmented habitats, may favour intra-pin outcrosses and increase mating opportunities in the population. A lower within-population structure can be expected because of a higher density of compatible mates and more random matings. Pin and thrum individuals showed similar levels of genetic variation. Three populations revealed a significant genetic differentiation between pin and thrum morphs ( F ST ⩾ 0.0 4 9 ), as a result of recent bottlenecks, spatial segregation of the morphs and/or disturbance of demographic processes. From our findings, we may hypothesize future changes in population mating processes in fragmented habitats, which might lead to pin excesses. However, due to slowing down life-history traits (long-lived perenniality and clonality), and demographic stochasticity randomly changing morph frequency, this may take time before being expressed. Evaluating the impact of morph frequencies on population dynamics is therefore essential for implementing conservation strategies of heterostylous plant species.

Importance of correlations among matrix entries in stochastic models in relation to number of transition matrices

Stochastic matrix models are used to predict population viability and the risk of extinction. Different stochastic methods require different amounts of estimation effort and may lead to divergent estimates. We used 16 transition matrices collected from ten populations of the perennial herb Primula veris to compare population estimates produced by different stochastic methods, such as selection of matrices, selection of vital rates, selection of matrix elements, and Tuljapurkar's approximation. Specifically, we tested the reliability of the methods using different numbers of transition matrices, and examined the importance of correlations among matrix entries. When correlations among matrix entries were included in the models, selection of vital rates produced the lowest and Tuljapurkar's approximation produced the highest estimates of mean population growth rates. Selection of matrices and matrix elements often produced nearly similar population estimates. Simulations based on incompletely estimated correlations among matrix entries considerably differed from those based on all correlations estimated, particularly when correlations were strong. The magnitude of correlations among matrix entries depended on the number of matrices, which made it difficult to generalize correlations within a species. Given that selection of vital rates or matrix elements is used, correlations among matrix entries should usually be included in the model, and they should preferably be estimated from the present data rather than according to other information of the species.