Nitrogen modulates strontium uptake and toxicity in Hypericum perforatum plants.

Strontium is an unavoidable element occurring in plants due to its abundance in the soil and similarity with calcium. To mimic natural conditions, impacts of additional inorganic (nitrate) or organic (urea and allantoin) nitrogen sources (1 mM of each N form in addition to 3.53 mM N in the basic cultivation solution) or N deficit on strontium-induced changes (100 µM Sr) in the widely used medicinal plant Hypericum perforatum L. were studied. Though various effects of Sr on primary (stimulation of amino acids but depression of most Krebs acids, ascorbic acid and thiols) and secondary metabolites (stimulation of phenols but no change of pseudo/hypericin) or mineral elements were observed (reduction of Ca amount in both shoots and roots), organic N forms often mitigated negative action of Sr or even combined stimulatory impact was observed. Organic N forms also elevated shoot accumulation of Sr while N deficit reduced it. Additional N forms, rather than Sr itself, modulated reactive oxygen species and nitric oxide formation in the root tissue. Germination experiment showed no toxicity of Sr to H. perforatum up to 1 mM Sr and even stimulated accumulation of amino acids and phenols, indicating similar ontogenetic-related responses.

The opposite nitric oxide modulators do not lead to the opposite changes of metabolites under cadmium excess.

Responses of common medicinal herb Matricaria chamomilla to short-term cadmium (Cd) exposure (48 h) alone or in combination with nitric oxide (NO) scavenger (PTIO) or donor (SNP) were studied. Modulators revealed expected impact on NO formation (depletion under PTIO but elevation under SNP) and confirmed anomalous impact on Cd accumulation (stimulation by both modulators). Changes of proline and free amino acids in the roots and fatty acids (mainly α-linolenic acid) in the shoots indicate rather relation to elevated Cd uptake than to altered NO formation. On the contrary, root profile of fatty acids revealed the most prominent changes in response to PTIO (elevation of monounsaturated and polyunsaturated fatty acids) which confirm that NO depletion stimulates their biosynthesis. Soluble phenols were elevated by SNP co-application in the roots while accumulation of chlorogenic acid and umbelliferone revealed clear regulation by NO (i.e. enhancement in response to SNP and depletion in response to PTIO). Data indicate that quantities of some metabolites are affected by accumulated Cd or NO formation and that the opposite NO modulators do not automatically lead to the opposite changes of plant metabolites.

Metabolic Changes Induced by Silver Ions in Carlina acaulis.

Silver is one of the most toxic heavy metals for plants, inducing various toxic symptoms and metabolic changes. Here, the impact of Ag(I) on Carlina acaulis physiology and selected metabolites was studied using two Ag concentrations (1 or 10 µM) after 14 days of exposure. The higher concentration of Ag(I) evoked reduction of growth, while 1 µM Ag had a growth-promoting effect on root biomass. The translocation factor (<0.04) showed that Ag was mainly retained in the roots. The 1 µM Ag concentration increased the level of low-molecular-weight organic acids (LMWOAs), while 10 µM Ag depleted these compounds in the roots. The increased concentration of Ag(I) elevated the accumulation of phytochelatins (PCs) in the roots and reduced glutathione (GSH) in the shoots (but not in the roots). At 1 µM, Ag(I) elevated the level of phenolic and triterpene acids, while the 10 µM Ag treatment increased the carlina oxide content in the roots. The obtained results indicate an alteration of metabolic pathways of C. acaulis to cope with different levels of Ag(I) stress. Our data imply that the intracellular binding of Ag(I) and nonenzymatic antioxidants contribute to the protection against low concentrations of Ag ions.

Applicability of a Monolithic Column for Separation of Isoquinoline Alkalodis from Chelidonium majus Extract.

Isoquinoline alkaloids are the main group of secondary metabolites present in Chelidonium majus extracts, and they are still the object of interest of many researchers. Therefore, the development of methods for the investigation and separation of the alkaloids is still an important task. In this work, the application potential of a silica-based monolithic column for the separation of alkaloids was assessed. The influence of the organic modifier, temperature, salt concentration, and pH of the eluent on basic chromatographic parameters such as retention, resolution between neighboring peaks, chromatographic plate numbers, and peak asymmetry were investigated. Based on the obtained results, a gradient elution program was developed and used to separate and quantitatively determine the main alkaloids in a Chelidonium majus root extract.

The Impact of Long-and Short-Term Strontium Treatment on Metabolites and Minerals in Glycine max.

The impact of long-term exposure to Sr2+ (LTE, four doses, 43.5 mg Sr2+ per pot, with a total of 174 mg Sr2+ per pot during the entire period of cultivation) and short-term exposure to Sr2+ (STE, one dose, 870 mg Sr2+ per pot four days before harvest) on the content of phytoestrogens and allantoin in soybeans were compared. Sr2+ accumulation, the effect on the concentration of macroelements, and basic physiology were also analyzed. LTE reduced the content of malonyldaidzin and malonylgenistin in the roots (58% and 50% compared to the control, respectively). STE increased the amount of all isoflavones in the stem and genistein in the leaves and decreased the content of malonyldaidzin and malonylgenistin in the leaves (55% and 48% compared to the control, respectively) and roots (69% and 62% of the control, respectively) as well as genistein and coumestrol in the roots (both 50% compared to the control). Sr2+ presence stimulated the accumulation of allantoin in the roots (three-fold higher than in the control), but only STE had similar effects on the shoots. In contrast to LTE, Sr2+ was transported extensively from the roots to the leaves under STE. In comparison to the control, LTE resulted in an increase in the Ca content in the stem by 36%, whereas Ca2+ accumulation in the leaves, stems, and roots increased by 60%, 80%, and 36%, respectively, under STE. Additionally, a significant accumulation of K was found only in the roots of the LTE group. The chlorophyll content did not differ between the treatments. Overall, the production of phytoestrogens and Sr accumulation were affected by both the applied dose and the duration of exposure to Sr.

Effect of Diosmin Administration in Patients with Chronic Venous Disorders on Selected Factors Affecting Angiogenesis.

Diosmin is a natural compound with a wide range of biological activity, e.g., it improves lymphatic drainage, supports microcirculation, and increases venous tone, and venous elasticity, hence, it is applied in the pharmacotherapy of chronic venous disorders (CVD). The aim of this study was to assess the correlation between diosmin administration (2 × 600 mg daily) in patients suffering from CVD and the levels of selected factors influencing angiogenesis, which are involved in CVD pathophysiology. Thirty-five CVD patients were examined. Levels of plasma tumor necrosis factor alpha (TNF alpha), vascular endothelial growth factor (VEGF-A and VEGF-C); angiostatin, interleukin 6 (IL-6), fibroblast growth factor 2 (FGF2); and plasminogen (PLG) were measured with an Elisa assay before and after three months of diosmin administration. The clinical symptoms of CVD were monitored using ultrasound images, echo Doppler assay, visual analogue scale (VAS), and measurement of the leg circumference. The average content of TNF alpha, VEGF-C, VEGF-A IL-6, and FGF2 decreased after the therapy with diosmin in a significant manner; with p < 0.001, p < 0.05, p < 0.05, p < 0.01, and p < 0.01, respectively, and a significant (p < 0.05) increase in the plasma angiostatin level after the three-month treatment was found. A significant (p < 0.05) decrease in edema and the average leg circumference of the patients was observed after the therapy. Diosmin influences the angiogenic and inflammatory mechanisms involved in the pathophysiology of edema presented in patients with a different class of CVD.

The Activity of Isoquinoline Alkaloids and Extracts from Chelidonium majus against Pathogenic Bacteria and Candida sp.

Chelidonium majus (Papaveraceae) extracts exhibit antimicrobial activity due to the complex alkaloid composition. The aim of the research was to evaluate the antimicrobial potential of extracts from wild plants and in vitro cultures, as well as seven major individual alkaloids. Plant material derived from different natural habitats and in vitro cultures was used for the phytochemical analysis and antimicrobial tests. The composition of alkaloids was analyzed using chromatographic techniques (HPLC with DAD detection). The results have shown that roots contained higher number and amounts of alkaloids in comparison to aerial parts. All tested plant extracts manifested antimicrobial activity, related to different chemical structures of the alkaloids. Root extract used at 31.25-62.5 mg/L strongly reduced bacterial biomass. From the seven individually tested alkaloids, chelerythrine was the most effective against P. aeruginosa (MIC at 1.9 mg/L), while sanguinarine against S. aureus (MIC at 1.9 mg/L). Strong antifungal activity was observed against C. albicans when chelerythrine, chelidonine, and aerial parts extract were used. The experiments with plant extracts, individually tested alkaloids, and variable combinations of the latter allowed for a deeper insight into the potential mechanisms affecting the activity of this group of compounds.

Historical and traditional medical applications of Carlina acaulis L. - A critical ethnopharmacological review.

ETHNOPHARMACOLOGICAL RELEVANCE: The genus Carlina (Asteraceae) comprises approx. 30 plant species growing in Europe and Asia. Carlina acaulis L. was widely used as medicine, especially in Germany and Poland. The plant root was applied e.g. to treat various skin diseases and as a diaphoretic, diuretic, and anthelmintic agent. At the end of the 19th century, the medicinal use of the plant ceased though C. acaulis roots are still used in folk medicine, especially in rural areas of the Balkans where the plant grows in the wild. AIM OF THE REVIEW: The aim of this review is to compile historical and current ethnomedical uses of C. acaulis and compare these applications with the results of modern chemical and pharmacological research of this plant. This analysis may give a lead for further research on this old, almost forgotten medicinal plant. METHODOLOGY: Historical data on the medicinal application of C. acaulis over many centuries were collected. The historical uses were compared with the contemporary indications. The present day knowledge on the chemistry and biological activity of C. acaulis is also reviewed. This information is compared with the ethnopharmacological information. RESULTS: Plants from the genus Carlina were already described by Theophrastus of Eresus, Dioscorides, Pliny the Elder, and Galen of Pergamum. Carlinae radix was still featured in Renaissance botanical books and in official pharmacopoeias, but disappeared from the scene in the 19th century. The present day information on the phytochemistry and pharmacological activity of these plants is limited. There are some data on the occurrence of common plant ingredients such as essential oil, flavonoids, phenolic acids, triterpenes, inulin, and fatty acids. However, most concern the green parts of the plant. Its medicinal use remains poorly understood, as there are only very limited results of in-vitro studies on the roots. CONCLUSIONS: The historical uses of the plant can still be found in rural areas. Yet, detailed phytochemical and pharmacological research is needed to evaluate the effectiveness and safety of the medical use of Carlina acaulis.

Bacterial Isolate Inhabiting Spitsbergen Soil Modifies the Physiological Response of Phaseolus coccineus in Control Conditions and under Exogenous Application of Methyl Jasmonate and Copper Excess.

The aim of the study was to demonstrate the potential of the promotion and regulation of plant physiology and growth under control and copper stress conditions, and the impact of the exogenous application of methyl jasmonate on this potential. Runner bean plants were treated with methyl jasmonate (1 or 10 µM) (J; J1 or J10) and Cu (50 µM), and inoculated with a bacterial isolate (S17) originating from Spitsbergen soil, and identified as Pseudomonas luteola using the analytical profile index (API) test. Above- and under-ground plant parts were analyzed. The growth parameters; the concentration of the photosynthetic pigments, elements, flavonoids (FLAVO), phenolics (TPC), allantoin (ALLA), and low molecular weight organic acids (LMWOAs); the activity of antioxidant enzymes and enzymes of resistance induction pathways (e.g., superoxide dismutase (SOD), catalase (CAT), ascorbate (APX) and guaiacol (GPX) peroxidase, glucanase (GLU), and phenylalanine (PAL) and tyrosine ammonia-lyase (TAL)), and the antioxidant capacity (AC) were studied. The leaves exhibited substantially higher ALLA and LMWOA concentrations as well as PAL and TAL activities, whereas the roots mostly had higher activities for a majority of the enzymes tested (i.e., SOD, CAT, APX, GPX, and GLU). The inoculation with S17 mitigated the effect of the Cu stress. Under the Cu stress and in the presence of J10, isolate S17 caused an elevation of the shoot fresh weight, K concentration, and TAL activity in the leaves, and APX and GPX (also at J1) activities in the roots. In the absence of Cu, isolate S17 increased the root length and the shoot-to-root ratio, but without statistical significance. In these conditions, S17 contributed to a 236% and 34% enhancement of P and Mn, respectively, in the roots, and a 19% rise of N in the leaves. Under the Cu stress, S17 caused a significant increase in FLAVO and TPC in the leaves. Similarly, the levels of FLAVO, TPC, and AC were enhanced after inoculation with Cu and J1. Regardless of the presence of J, inoculation at Cu excess caused a reduction of SOD and CAT activities, and an elevation of GPX. The effects of inoculation were associated with the application of Cu and J, which modified plant response mainly in a concentration-dependent manner (e.g., PAL, TAL, and LMWOA levels). The conducted studies demonstrated the potential for isolate S17 in the promotion of plant growth.

Eleutherococcus Species Cultivated in Europe: A New Source of Compounds with Antiacetylcholinesterase, Antihyaluronidase, Anti-DPPH, and Cytotoxic Activities.

Secondary metabolites of the roots of Eleutherococcus spp. cultivated in Poland, or the bioactivity, are not fully known. The 75% methanol extracts of five Eleutherococcus spp. (E. senticosus, E. divaricatus, E. sessiliflorus, E. gracilistylus, and E. henryi) were examined for the content of polyphenols and phenolic acids as well as for antiacetylcholinesterase, antihyaluronidase, anti-DPPH∗, and cytotoxic activities. The richest in polyphenols were the roots of E. henryi (10.4 mg/g DW), while in flavonoids the roots of E. divaricatus (6.5 mg/g DW). The richest in phenolic acids occurred the roots of E. henryi [protocatechuic acid (1865 µg/g DE), caffeic acid (244 µg/g DE), and p-coumaric and ferulic acids (55 µg/g DE)]. The highest inhibition of AChE was observed for E. gracilistylus and E. sessiliflorus (32%), at the concentration of 100 µg/0.19 mL of the reaction mixture, while that of Hyal for the roots of E. henryi (40.7%), at the concentration of 100 µg/0.16 mL of the reaction mixture. Among five species tested, the E. henryi extract exhibited the strongest HL-60 cell line growth's inhibition (IC50 270 µg/mL). The extracts reduced DPPH∗ in a time-dependent mode, at the concentration of 0.8 mg/mL. After 90 min from 14.7 to 26.2%, DPPH∗was reduced. A phytochemical composition and activity of the Eleutherococcus species, cultivated in Poland, are still under research; however, on the basis of the results obtained, it may be concluded that they may become a source of phytochemicals and be useful for Europe's citizens.

Polyaniline based material as a new SPE sorbent for pre-treatment of Chelidonium majus extracts before chromatographic analysis of alkaloids.

In the present study, deprotonated and protonated polyaniline based sorbent obtained by in situ polymerization of aniline on silica (Si-PANI) was applied for SPE pretreatment of Chelidonium majus extracts before determination of benzophenanthridine, protoberberine and protopine alkaloids. The experimentally optimized conditions, such as protonated polyaniline impregnated with methanol as a sorbent, 2 mL of water and methanol mixture (1/1, v/v) as a washing solution, and 5 mL of 0.1 M methanolic solution of ammonia as an elution solvent, yielded the highest values of analyte recovery (above 96.5%) with simultaneous removal of undesirable plant matrix. Virtually modeled structure of polyaniline revealed possible occurrence of the mixed mode binding mechanism, based both on π-π and ion interactions on protonated form of polyaniline. Moreover, lack of significant changes of the polyaniline film quality assessed by Raman spectroscopy after series of ten experiments proved the reusability of the sorbent.

Allantoin attenuates cadmium-induced toxicity in cucumber plants.

Concentration-dependent responses of cucumber plants to cadmium (Cd, 5-15 µM) and/or allantoin (Alla, 10-1000 µM) have been investigated to detect a possible protective role of Alla under Cd excess. After 14 days of exposure, Alla often considerably reversed Cd-induced inhibition of growth and reduction of the content of photosynthetic pigments. Higher Alla doses depleted the Cd amount in shoots, which could be related to citric acid (increase in the shoots but depletion in the roots in Cd+Alla treatments) rather than to phytochelatins (Alla had a negative impact on the phytochelatin accumulation). An increase in the Alla concentration suppressed Cd-induced spatial H2O2 appearance, which does not seem to be related to antioxidative enzymes (low impact of Alla on catalase, ascorbate peroxidase, and guaiacol peroxidase). On the contrary, shoot glutathione and mainly ascorbic acid accumulation strongly increased in Cd+Alla treatments, indicating their prominent role in Alla-induced amelioration of Cd-stimulated oxidative stress and growth retardation. Similarly, phenolic metabolites (total soluble phenols and flavonols) were slightly influenced by Alla and their antioxidative action was not expected. We conclude that Alla-mediated attenuation of Cd-induced toxicity relies on enhanced accumulation of glutathione and ascorbate in the shoot tissue mainly, rather than on elevated antioxidative enzyme activities.

Methodological approach to determine carlina oxide - a main volatile constituent of Carlina acaulis L. essential oil.

In this work, a fast and low-cost voltammetric methodology for determination of carlina oxide in plant extracts was developed. The best results were obtained using a boron-doped diamond electrode (BDDE). The voltammetric measurements of carlina oxide were performed in a 0.1 mol/L solution of sulphuric acid. After 30 s of stirring the solution, differential pulse voltammograms (DPVs) were recorded from 0.5 to 1.8 V. The amplitude was 75 mV and the scan rate was 175 mV/s. Measurements were recorded in non-deaerated solutions. The background current was subtracted from each registered voltammogram; then they were cut from 0.5 to 1.5 V. The detection and quantification limits were 0.28 and 0.93 µg/L, respectively, and repeatability expressed as the relative standard deviation of 0.1 mg/L of carlina oxide was 1.9% (n = 5). The results were compared with those obtained using gas chromatography with a flame ionization detector and high performance liquid chromatography with a photodiode array detector.

The Impact of Different Cultivation Systems on the Content of Selected Secondary Metabolites and Antioxidant Activity of Carlina acaulis Plant Material.

Roots and leaves of Carlina acaulis L. are still used in ethnomedicine in many European countries; however, the limited occurrence of the plants and protection of this species necessitate a search for alternative ways for obtaining this plant material. In this study, in vitro cultures, hydroponic cultures, and field cultivation were applied to obtain the C. acaulis plant material. Its quality was evaluated using antioxidant activity tests and high performance liquid chromatography analysis. Our study showed that the antioxidant activity and the content of chlorogenic and 3,5-di-caffeoylquinic acid in roots of plants cultivated in hydroponics and field conditions were comparable. However, the amount of carlina oxide was significantly higher in plants from the field. The flavonoid content in leaves obtained from both cultivation systems was at the same level; however, the antioxidant activity and the content of the investigated metabolites were higher in the soil cultivation system. The callus line exhibited high differentiation in phytochemical compositions depending on the treatments and medium compositions.

Effect of Long-Term Strontium Exposure on the Content of Phytoestrogens and Allantoin in Soybean.

Abiotic stress, including metal excess, can modify plant metabolism. Here we investigated the influence of long-term strontium exposure (12 weeks, 0.5⁻4.0 mM Sr) on the content of phytoestrogens and allantoin as well as the mineral composition in soybean. Seven phytoestrogens were identified in the soybean: daidzin, glycitin, genistin, malonyldaidzin, malonylgenistin, daidzein, and coumestrol. The results showed that both malonyldaidzin and malonylgenistin were dominant phytoestrogens; however, the roots contained a relatively high amount of daidzein. It was found that strontium reduced the phytoestrogen content and decreased the antioxidant capacity. Strontium evoked depletion of the sum of all phytoestrogens by 40⁻70% in the leaves, 25⁻50% in the stems and in the seeds, depending on the strontium concentration. In the roots, 0.5 and 4.0 mM of strontium decreased the total phytoestrogen content by 25 and 55%, respectively, while 2.0 mM of strontium did not exert an effect on their accumulation. On the other hand, strontium ions induced allantoin accumulation mainly in the roots. Strontium was preferentially accumulated in the leaves, with a slight impact on macro- and micro-nutrients. Our research showed strontium-secondary metabolites interaction in the soybean, which can be useful for obtaining a natural pharmaceutical product containing both strontium and phytoestrogens for remediation of postmenopausal osteoporosis.

Metabolic changes induced by manganese in chamomile.

Manganese (Mn) uptake and toxicity in chamomile (Matricaria chamomilla) and changes of phenolic metabolites in plants grown in the soil (1000 µM Mn2+) or hydroponic culture (100 or 1000 µM Mn2+) were studied. Under soil cultivation, Mn excess reduced growth and induced symptoms of oxidative stress (including total ROS, hydroxyl radical and lipid peroxidation as detected by fluorescence microscopy), concomitantly with depletion of non-protein thiols and ascorbic acid. Total soluble phenols and individual phenolic acids were rather depleted (p-coumaric, chlorogenic, and protocatechuic acids) or unaltered (vanillic and caffeic acids). Shoot Mn content reached 2806 µg/g DW with BAF 51.0 in the soil culture. In hydroponics, tetraploid plants contained less Mn in both shoots and roots than diploid ones with bioaccumulation factor and translocation factor (diploid/tetraploid) 57.1/37.9 and 0.39/0.32 in 1000 µM Mn treatment. Plants cultured in hydroponics revealed stimulation of some phenolic acids, mainly chlorogenic acid in the shoots and p-hydroxybenzoic and vanillic acids in the roots (more extensively in tetraploid ones which contained less Mn). Data indicate that excessive Mn accumulation has negative impact not only on the growth but also on phenolic metabolites in young plants mainly. Detailed comparison of the observed metabolic changes with limited literature focused on Mn physiology is provided as well.

Influence of Diosmin Treatment on the Level of Oxidative Stress Markers in Patients with Chronic Venous Insufficiency.

Oxidative stress plays an important role in the pathophysiology of many human disorders, while antioxidants prevent the development of various adverse symptoms. Diosmin is a natural flavonoid applied in vascular system disorders, especially in chronic venous insufficiency (CVI), and it plays a significant part in the alleviation of CVI symptoms. Due to antioxidant activity, it also has the ability to scavenge the oxygen free radicals and hence decreases the level of oxidative stress biomarkers, such as prostaglandins and their precursors-isoprostanes. In the study, the influence of diosmin treatment on the level of isoprostanes in plasma samples of patients suffering from CVI was examined. The qualitative analysis was performed using high-performance liquid chromatography with spectrometry detection (LC-MS). The statistically significant decrease of isoprostane content after 3 months of treatment was observed within the studied group; however, the most significant changes were observed in patients who smoke.

Greater Celandine's Ups and Downs-21 Centuries of Medicinal Uses of Chelidonium majus From the Viewpoint of Today's Pharmacology.

As antique as Dioscorides era are the first records on using Chelidonium as a remedy to several sicknesses. Inspired by the "signatura rerum" principle and an apparent ancient folk tradition, various indications were given, such as anti-jaundice and cholagogue, pain-relieving, and quite often mentioned-ophthalmological problems. Central and Eastern European folk medicine has always been using this herb extensively. In this region, the plant is known under many unique vernacular names, especially in Slavonic languages, associated or not with old Greek relation to "chelidon"-the swallow. Typically for Papaveroidae subfamily, yellow-colored latex is produced in abundance and leaks intensely upon injury. Major pharmacologically relevant components, most of which were first isolated over a century ago, are isoquinoline alkaloids-berberine, chelerythrine, chelidonine, coptisine, sanguinarine. Modern pharmacology took interest in this herb but it has not ended up in gaining an officially approved and evidence-based herbal medicine status. On the contrary, the number of relevant studies and publications tended to drop. Recently, some controversial reports and sometimes insufficiently proven studies appeared, suggesting anticancer properties. Anticancer potential was in line with anecdotical knowledge spread in East European countries, however, in the absence of directly-acting cytostatic compounds, some other mechanisms might be involved. Other properties that could boost the interest in this herb are antimicrobial and antiviral activities. Being a common synanthropic weed or ruderal plant, C. majus spreads in all temperate Eurasia and acclimates well to North America. Little is known about the natural variation of bioactive metabolites, including several aforementioned isoquinoline alkaloids. In this review, we put together older and recent literature data on phytochemistry, pharmacology, and clinical studies on C. majus aiming at a critical evaluation of state-of-the-art from the viewpoint of historical and folk indications. The controversies around this herb, the safety and drug quality issues and a prospective role in phytotherapy are discussed as well.

Systematic Evaluation of Chromatographic Parameters for Isoquinoline Alkaloids on XB-C18 Core-Shell Column Using Different Mobile Phase Compositions.

Chelidonium majus L. is a rich source of isoquinoline alkaloids with confirmed anti-inflammatory, choleretic, spasmolytic, antitumor, and antimicrobial activities. However, their chromatographic analysis is difficult because they may exist both in charged and uncharged forms and may result in the irregular peak shape and the decrease in chromatographic system efficacy. In the present work, the separation of main C. majus alkaloids was optimized using a new-generation XB-C18 endcapped core-shell column dedicated for analysis of alkaline compounds. The influence of organic modifier concentration, addition of salts, and pH of eluents on chromatographic parameters such as retention, resolution, chromatographic plate numbers, and peak asymmetry was investigated. The results were applied to elaborate the optimal chromatographic system for simultaneous quantification of seven alkaloids from the root, herb, and fruit of C. majus.

Methodological aspects of biologically active compounds quantification in the genus Hypericum.

Accumulation of selected secondary metabolites in two Hypericum species (H. perforatum and H. annulatum) was compared in their vegetative parts (stems and leaves) and in terms of the extraction solvent (80% aq. methanol or 60% aq. ethanol). The presence of chlorogenic acid and quercitrin was not detected in stem of both species. Almost all metabolites were more accumulated in the leaves than in the stems (rutin, hyperoside, quercetin and hypericin) but epicatechin showed the opposite in both species and hyperforin in H. annulatum. Extraction solvents showed rather species-specific differences with EtOH being more suitable for the extraction of hypericin, quercetin, quercitrin, and hyperoside (on average, for both the leaves and stems, extraction increased by approximately 130, 30, 25, and 15%, respectively) while MeOH for the extraction of epicatechin, rutin, and hyperforin (increased extraction by approximately 50, 40, and 35%, respectively). On the other hand, content of total soluble phenols did not differ in relation to solvent in any organ or species. Various ages of H. annulatum plants did not show dramatic impact on the amount of metabolites. Subsequently, the usefulness of capillary electrophoresis (CE) as an alternative to HPLC for the quantification of metabolites in H. perforatum was tested and results showed non-significant differences between CE and HPLC with the methods we developed (the difference did not exceed 10%).