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The subject of this study is the composition of low-molecular-weight metabolites in downy birch (Betula pubescens) buds and their participation in protection from various kinds of stress. Using the GC-MS, 640 compounds were detected, of which 314 were identified in downy birch buds for the first time. The volatile components detected using the SPME technique mainly consisted (about 70% of the total ionic current of the chromatogram, TIC) of mixtures of sesquiterpenoids. The exudate covering the buds, along with sesquiterpenoids (approximately 60% of TIC), included flavonoids (25% of TIC). The main part of the material extracted by supercritical carbon dioxide from buds comprised sesquiterpenoids and triterpenoids (47 and 28% of TIC, respectively). Via column chromatography, 25 known compounds (mainly flavonoids and triterpenoids) were isolated, most of which were first discovered in the buds of downy birch. Many compounds of these classes have strong biological activity and probably either directly or indirectly perform a protective function in birch buds. An assumption is made about the biological role of a number of secondary metabolites (such as volatile isomeric megastigmatriens and triterpene seco-acids) as well as about these compounds’ possible means of biosynthesis, which were first discovered in the buds of downy birch.

The aim of the present work was to investigate and compare the content of phenolic compounds in four taxa of Erigeron acris L. s. l.: E. acris (EAA), E. acris subsp. droebachiensis (O.F. Müll.) Arcang. (EAD), E. acris subsp. serotinus (Weihe) Greuter (EAS) and E. ×huelsenii Vatke (EH), a hybrid between E. acris and E. canadensis L. The total flavonoid content was determined by Christ-Müller method and the total phenolic acid content was determined by the method utilizing Arnov’s reagent. The method using ultra high performance liquid chromatography with photodiode array detection (UHPLC-PDA) was applied for the separation, identification and quantification of nine phenolic compounds (protocatechuic acid, chlorogenic acid, caffeic acid, 6′-O-caffeoylerigeroside, scutellarein-7-O-β-D-glucuronide, quercetin 3-O-glucoside, 4,5-dicaffeoylquinic acid, quercetin and luteolin) in the aerial parts of E. acris s. l. The chromatographic separation was carried out using a BEH C18 column packed with 1.7-μm particles and gradient elution with a mobile phase of water and methanol, both containing 0.02% (v/v) trifluoroacetic acid. The four investigated taxa of E. acris s. l. differed in the composition and the content of phenolic compounds. The main substances determined in the methanolic herbal extracts were: scutellarein-7-O-β-D-glucuronide (EAA, EAS, EAD and EH), 6′-O-caffeoylerigeroside (EAA, EAD and EH) and chlorogenic acid (EAS and EH). Moreover, the results indicated that five of the nine tested compounds were found in all investigated extracts from herbs of E. acris s. l. Two of them (6′-O-caffeoylerigeroside and scutellarein-7-O-β-D-glucuronide) could be selected as potential chemotaxonomic markers of the genus Erigeron L.

Rosmarinic acid (RA) is a natural phenylpropanoid with numerous pharmacological activities. Because of limited studies of the effects of RA action in gastric cancer cells we examined how 100 and 200 μM acid influences MMPs, TIMPs, collagen, MUC1 and specific sugar antigens in gastric adenocarcinoma CRL-1739 cells. We revealed inhibitory effect of RA on MMP-9 activity what was correlated with increased collagen type I expression, main ECM substrate degraded by MMPs. Tissue inhibitor of MMPs, TIMP-1 but not TIMP-2 was significantly decreased on the protein level and increased on mRNA level by RA action what can suggest TIMP-1 independent inhibitory action of an acid on MMP-9 activity. Glycosylation of gastric cancer proteins was also effected by RA. ELISA tests revealed inhibitory effect of an acid on Tn antigen in cell lysates and culture supernatant and on T antigen in cell lysates. RA inhibited also sialylated Tn antigen in protein of culture supernatant and sialyl T in cell lysates. Extracellular domain of MUC1 mucin, main carrier of Tn and T antigens was significantly inhibited by higher dose of RA. The data suggest potential usefulness of RA as a complementary agent supporting chemotherapy in cancer treatment.

Flavonoid compound scutellarin (Scu) is quite frequently met in the plant kingdom, particularly in the genus Scutellaria (Lamiaceae) and Erigeron (Asteraceae). The extract of the herb of Erigeron breviscapus, containing this component in high amount, has been used for many years in traditional Chinese medicine. In recent years, studies have made great progress on the usefulness of Scu for treating various diseases by testing its mechanism of action. They support the traditional use of Scu rich plant in heart and cerebral ischemia. Scu can potentially be applied in Alzheimer’s disease, Helicobacter pylori infection, vascular complications of diabetes and as an inhibitor of certain carcinomas. Various methods were designed to improve its isolation from plant material, solubility, absorption and bioavailability. On the basis of recent studies, it is suggested that Scu could be a promising candidate for new natural drug and deserves particular attention in further research and development.

CK2 is a pleiotropic, constitutively active protein kinase responsible for the phosphorylation of more than 300 physiological substrates. Typically, this enzyme is found in tetrameric form consisting of two regulatory subunits CK2β and two catalytic subunits CK2α or CK2α′. Several natural occurring flavonoids were tested for their ability to inhibit both CK2 holoenzymes, CK2α2β2 and CK2α′2β2. We identified few substances selectively inhibiting only the α′ subunit. Other compounds showed similar effect towards all four isoforms. In some cases, like chrysoeriol, pedalitin, apigenin, and luteolin, the α2β2 holoenzyme was at least six times better inhibited than the free α subunit. Otherwise, we have found a luteolin derivative decreased the kinase activity of CK2α′ with an IC50 value of 0.8 μM, but the holoenzyme only with 9.5 µM.

The article presents for the first time the linear temperature programmed retention indices on a column with stationary phases of 5% phenylpolydimethyl silicone and the mass spectra of trimethylsilyl (TMS) derivatives of 71 glycosides (both commercial preparations and compounds extracted from plant tissues) which were not characterized earlier by these parameters. Converted to their TMS derivatives, the glycosides were thermally stable: they exhibited single peaks on their chromatograms without products of thermal decomposition. Therefore this work demonstrates the suitability of high resolution-high temperature gas chromatography (HR-HT/GC) to analyse different groups of glycosides including compounds with disaccharide moieties without the necessity of their hydrolyses. Since a limited number of commercial and plant-isolated glycosides were available, an attempt was made to assess their retention indices using the known "structure-retention relationships" approach. It was demonstrated that the retention indices of silanised glycosides and their aglycones were characterized by a linear dependence.

The present study investigated the magnitude and mechanism of the cytotoxic effect on selected cancer cell lines of 3,4-seco-urs-4(23),20(30)-dien-3-oic acid (1), 3,4-seco-olean-4(24)-en-19-oxo-3-oic acid (2), and 3,4-seco-urs-4(23),20(30)-dien-19-ol-3-oic acid (3) isolated from downy birch (Betula pubescens) buds by carbon dioxide supercritical fluid extraction and gradient column chromatography. Cell viability in six human cancer lines exposed to these compounds was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Apoptosis was quantified by annexin V/propidium iodide staining of gastric cancer AGS and colorectal cancer DLD-1 cells. To evaluate the mechanism of apoptosis, the expression of apoptosis-related proteins was analyzed by Western blot. Compound 1 exhibited non-specific toxicity, while compounds 2 and 3 were specifically toxic to colon and stomach cancer cells. The toxicity of compounds 2 and 3 against these two cell lines was greater than for compound 1. Cleavage of caspase-8, -9, and -3 was found in AGS and DLD-1 cells treated with all three seco-acids, indicating the induction of apoptosis via extrinsic and intrinsic pathways. Therefore, triterpene seco-acids (1&ndash, 3) decreased cell viability by apoptosis induction. AGS and DLD-1 cells were more susceptible to seco-acids with an oxidized C19 than normal fibroblasts. Hence, it made them a new group of triterpenes with potential anticancer activity.

Introduction The quality of herbs is directly related to the presence of polyphenolic antioxidants. This is the first report on the quantification of individual polyphenolic constituents of Erigeron acris L. Objective To develop a new method using ultra-high performance liquid chromatography with photodiode array and chemiluminescence (UHPLC-PDA-CL) detection for the separation and determination of polyphenols in Erigeron acris extracts. Methodology The methanolic extracts from leaves and inflorescences of Erigeron acris were prepared by ultrasound assisted extraction. The chromatographic separation was performed on C18 column packed with 1.7-μm particles. The post-column CL detection was based on the enhancing effect of polyphenols on the CL generated in manganese(IV)–hexametaphosphate–formaldehyde system. Results The UHPLC method allowed to separate polyphenols in a short running time (13 min), which was three times shorter compared with traditional HPLC. The CL detection was characterised by 6–48 times higher sensitivity and up to three times lower detection limits compared to PDA detection. Qualitative and quantitative differences were observed in polyphenolic composition of Erigeron acris extracts. The main components of leaves were scutellarin and chlorogenic acid, whereas in inflorescences quercetin 3-O-glucoside was predominant. Conclusion Coupling of UHPLC with CL detection has been developed for the first time. This advanced chromatographic technique coupled with sensitive CL detection is a powerful approach for the investigation of polyphenolic profiles in natural products. The shorter analysis time and diminished waste generation makes the UHPLC method more environmentally friendly and more cost-effective in comparison with conventional HPLC. Copyright © 2016 John Wiley & Sons, Ltd.

The mixture of three phytosterols (campesterol, stigmasterol and β-sitosterol), β-sitosterol 3-O-glucoside and syringin were isolated from hexane and methanol extract of Cirsium rivulare roots after chromatographic separation. The main component of the source was syringin which was obtained with the yield 0.08% of the dry source. In hexane extract, the qualitative and quantitative composition of fatty acids was determined. The predominant component was linoleic acid (23.31 mg/g of extract). The extracts showed antioxidant activity. The ability to scavenge DPPH• free radical was in correlation with appointed total phenol content. The not-defatted methanolic extract was the most active. Hexane and defatted methanol extracts showed moderate antibacterial activity against G(+) and G(−) strains with MIC and MBC ranged from 25 to 200 μg/mL.

Birch buds (Gemmae Betulae) are widely used in Russian and Chinese traditional medicine mainly as a diuretic and diaphoretic agent but also as an antiseptic, anti-inflammatory and analgesic. Despite the long history of therapeutic use of birch buds in folk medicine, the existing information on their chemical composition and pharmacological effects is insufficient. This circumstance warrants further study of the chemistry and pharmacology of birch buds. The present study was designed to investigate (a) the chemical composition of buds from two species of white birch and (b) the in vitro cytotoxic effect of extracts from these sources on selected tumour cells. Extracts from Betula pubescens Ehrh. and Betula pendula Roth. buds were obtained using three different methods: carbon dioxide supercritical fluid extraction (SFE), washing of exudate covering whole buds, and extraction of milled buds with diethyl ether. The chemical composition of extracts was investigated by GC-MS. Cytotoxicity was determined by MTT assay, and cell proliferation was determined by [3H]thymidine uptake in cancer cells and normal skin fibroblasts. The GC-MS investigation identified a total of 150 substances of different classes. The chemical composition of B. pubescens and B. pendula buds differed, with bud extracts from the former containing a relatively high quantity of sesquiterpenoids and flavonoids, while the main components of extracts from the latter were triterpenoids. The results of the biological assay indicated that birch bud extracts demonstrated time- and concentration-dependent and differential cytotoxicity. The highest cytotoxic activity demonstrated bud exudates and SFE extracts obtained from both Betula species. The rich chemical composition of birch buds suggests the possibility of a wider spectrum of biological activity than previously thought. Birch bud extracts could be a promising source of compounds with cytotoxic activity against various cancers.

Purpose In aging skin and some skin disorders, components of skin extracellular matrix (ECM) are disturbed and therefore research to find skin drugs is important. Evaluation of anethole impact on collagen, GAGs and MMP-2 in human skin fibroblasts was the aim of this study. Materials and methods For collagen assay the Sircol dye, 5-[3H]proline and real time-PCR were used. MMP-2 activity was detected by zymography. GAG concentration was determined using 1,9-dimethylmethylene blue (DMMB). Cell viability was assayed with MTT. Results In cells treated with 1 and 10 μM anethole, a significant increase in collagen synthesis was demonstrated. In contrast, collagen synthesis was significantly decreased in cells exposed to 100 μM anethole. Similar alterations were found in collagen type I expression. The concentration of collagen secreted into the medium was higher only in cells exposed to 1 μM anethole, while it was lower under the influence of higher compound concentrations. It may be due to the lack of pro-MMP-2 activation at 1 μM and a significant increase in the level of MMP-2 at 10 and 100 μM anethole. GAG concentration was reduced under the influence of 100 μM anethole, whereas anethole at lower concentrations revealed the ability to prevent H2O2-induced GAG increase. No significant cytotoxicity of anethole to fibroblasts was noted. Conclusions Our findings demonstrate the concentration-dependent action of anethole on the crucial components of ECM in cultured skin fibroblasts, which may be somewhat beneficial and may possibly be developed towards a therapeutic use in some skin disorders.

Matricaria is a widespread genus of flowering plants of the family Asteraceae that grow in temperate regions of Europe, Asia, America and Africa. Some of the species are also naturalized in Australia. Some species of this genus such as Chamomiles are recognized medicinal plants and cultivated in several countries for commercial purposes: to obtain its blue essence, as herbal tea, and for pharmaceutical or cosmeceutical uses. The phytochemical composition of Matricaria spp. includes volatile terpenoids (e.g., α-bisabolol, bisabolol oxide A and B, β-trans-farnesene and chamazulene), sesquiterpene lactones such as matricin, and phenolic compounds (flavonoids, coumarins and phenolic acids). Their essential oil is obtained from the fresh or dried inflorescences by steam distillation, and additionally cohobation of the remaining water. The volatile composition of the essential oil, especially the content of the valuable components α-bisabolol and chamazulene, depends on the plant part, origin and quality of the source, genetic, and environmental factors. Moreover, other parameters, such as season of harvest and methods of extraction, can affect the extraction yield of the essential oils/extracts, their composition and, therefore, their bioactivity. Due to the importance of this genus and particularly M. recutita (M. chamomilla), this review focus on its cultivation, factor affecting essential oils’ composition and their role in traditional medicine, as antibacterial agents and finally as food preservatives.

Novel methods for the determination of polyphenolic antioxidants present in extracts from inflorescences of Cirsium vulgare (Savi) Ten. based on ultra high performance liquid chromatography with photodiode array and chemiluminescence detection have been developed. Under the optimized conditions of chromatographic separation the analytical characteristic of the method was performed. The proposed method was successfully applied to the determination of ten polyphenols present in inflorescences of Cirsium vulgare. A comparison of the contents of analytes in extracts prepared by using various extraction media (methanol, ethanol, 70% methanol, 70% ethanol and water) was carried out for the first time. For the post-column detection of scavenging activity of polyphenolic antioxidants against reactive oxygen species (H2O2, •OH, O2• −) three systems based on chemiluminescence of luminol were used. A review of the current scientific literature shows that this is the first report on the application of luminol-based post-column detection for the on-line investigation of •OH scavenging activity. The main compound determined in extracts from inflorescences of Cirsium vulgare was apigenin 7-O-glucuronide, whereas the highest antioxidant activity was observed for chlorogenic acid, luteolin 7-O-glucoside and apigenin. This article is protected by copyright. All rights reserved

It was found that weak chemiluminescence of manganese(IV)-hexametaphosphate-formaldehyde system was greatly enhanced by plant polyphenolic compounds. Based on this finding, a new flow injection chemiluminescence method (FI-CL) was developed for the determination of the total content of polyphenols in plant extracts. The calibration graph obtained for standard solutions of 6-hydroxyluteolin 7-O-glucoside (6OHLG) was linear in the range 0.001-0.8 μg mL(-1). The method was simple, rapid (203 samples h(-1)) and sensitive with a detection limit of 0.25 ng mL(-1). The FI-CL method was successfully applied to the determination of the total polyphenols content (as 6OHLG equivalents) in methanolic, ethyl acetate and aqueous extracts from leaves of Cirsium palustre (L.). Two types of solvent extraction methods (reflux and ultrasound assisted extraction) were used and compared in terms of extraction efficiency. A positive, significant linear correlation between the results obtained by FI-CL method and spectrophotometric methods was observed.

Ten flavonoids belonging to the subclasses of flavones, flavanones and aurones were isolated from methanolic extract of Cirsium palustre leaves after multistep chromatographic separation. Their structures were elucidated with spectroscopic methods. All compounds, except for luteolin 7-O-glucoside, were isolated for the first time. Four compounds—eriodictyol 7-O-glucoside (6), 6-hydroxyluteolin 7-O-glucoside (11), scutellarein 7-O-glucoside (12) and pedalitin (14)—were tested for their effect on collagen expression in normal human dermal fibroblasts. Among them, compound 11 at 40 μM and compound 14, at all concentrations used (1, 20, 40 μM), significantly enhanced the level of total collagen secreted into the medium. Furthermore, compound 11 significantly stimulated type I collagen expression, whereas compound 14 activated type I and III collagen expression at the mRNA level, depending on concentration. MMP-2 activity was inhibited by all study compounds, with the greatest effect recorded with compound 14 at 20 μM. The lack of effect on collagen content in the medium of compound 6- and compound 12-treated cells, besides an increase in COL1A1 and COL1A2 expression, might be caused by diminished expression of HSP47 gene, resulting in decreased procollagen secretion. Future study of compounds 11 and 14 for their potential therapeutic use in conditions connected with collagen biosynthesis deficiency is required. Copyright © 2014 John Wiley & Sons, Ltd.

CK2 is a ubiquitous protein kinase involved in many cell functions. During the last years it became an interesting target in cancer research. A series of flavonoid compounds was tested as inhibitors of protein kinase CK2. Several substances were found to be highly active against both catalytic subunits with IC 50 values below 1 μM in case of CK2α′. The most promising inhibitor we identified is chrysoeriol with IC 50 values of 250 and 34 nM for CK2α and CK2α′, respectively.

In this study, antimicrobial properties and toxicity of extracts from Cirsium spp.: Cirsium arvense, C. oleraceum, C. palustre, C. rivulare and C. vulgare in combination with sodium picolinate (PS) or sodium benzoate (BS), were investigated. Three micro-organisms were used: Staphylococcus aureus, Bacillus subtilis and Pseudomonas aeruginosa. Minimum inhibitory concentration (MIC) of extracts was found at 1.56-50.0 mg mL(-1). Unlike the case of BS, adding PS to extracts from flowers of C. palustre and C. arvense enhanced their antimicrobial effect on S. aureus (MIC from 6.25-12.5 mg mL(-1) to 1.25-5.0 mg mL(-1)). An MTT test was used to study toxicity effects. The extracts from C. palustre or C. arvense mixed with PS had a concentration-dependent, slightly cytotoxic or stimulating effect on the viability of normal human skin fibroblasts. The total phenolic content (TPC) of samples varied from 44 to 178 mg gallic acid equivalent per 1 g of extract. The highest TPC was observed in C. palustre (l) and C. oleraceum (f). Our results did not show any correlation between antimicrobial activities and TPC. Cirsium palustre (f) and C. arvense (f) extracts were analysed by gas chromatography/mass spectrometry (GC/MS). About 30 compounds were found to be present in extracts from two Cirsium species in amounts of not less than 0.2% of TIC.

The chemical compositions of essential oils from the roots of Erigeron acris and Erigeron annuus were studied. The essential oils were obtained by hydrodistillation in 1.0% and 0.05% yield, respectively, and analyzed by GC, GC-MS. Fifty four and forty seven constituents were identified. Predominant constituents of both oils were poly-acetylene esters: (Z,Z)-matricaria ester (49.4% and 45.9%, respectively) and (Z)-lachnophyllum ester (37.2% and 27.5%, respectively), that were accompanied by their stereoisomers as well as appropriate lactones. Polyacetylenic compounds amounted to 92.1% of E. acris oil and 85.8% of E. annuus oil. Both oils contained the same monoterpene hydrocarbons, amounting to 4.2% and 5.8%, respectively, and traces of almost the same monoterpene oxygenated compounds. The dominant sesquiterpenes in E. acris were elemenes and tricyclic sesquiterpene hydrocarbons, while in E. annuus beta-sesquiphellandrene and beta-bisabolene dominated. After flash chromatography of essential oil from E. acris, fractions contained acetylene esters and acetylene lactones were obtained. The configuration about double bonds for these compounds has been elucidated on the basis of (1)H- and (13)C-NMR analysis.

The essential oil from flowers, leaves, stems and roots of Erigeron annuus (L.) Pers. was analyzed by GC, GC-MS and 1H-NMR spectroscopy. More than 60 compounds were identified. Germacrene D (47.2%) and (Z)-lachnophyllum ester (10.2%) were the main constituents of the flower oil. The leaf oil and stem oil contained mainly germacrene D (55.6% and 44.5% respectively) and β-caryophyllene (9.4% and 13.4%), whereas in the root oil dominated matricaria ester (59.9%) and (Z)-lachnophyllum ester (34.9%).

A new flavonoid compound isokaempferide 7-O-beta-d-(6''-methylglucuronide) (1) was isolated from flowers of Cirsium rivulare (Jacq.) All. Its structure was elucidated on the basis of spectroscopic studies (UV, (1)H and (13)C NMR, FABMS). Moreover, five known flavonoid compounds: apigenin 7-(6''-methylglucuronide) (2), hispidulin 7-glucoside (3), isokaempferide 7-glucuronide (4), kaempferol 3-galactoside (5), pectolinarin (6) were isolated from this source for the first time. The antimicrobial activity of aqueous, methanolic and 70% ethanolic extracts of Cirsium rivulare flowers and leaves was also evaluated.

In the hexane extract of C. palustre and C. rivulare fruits, fatty acids, sterols, triterpenes, and volatile compounds were analyzed by the GC-MS-FID method. In the methanolic extracts, total phenol content was estimated. The antioxidant activity of both extracts was measured with DPPH assay and expressed in % scavenged DPPH.

The first method for the simultaneous determination of polyphenolic antioxidants in extracts from leaves of Cirsium palustre based on high performance liquid chromatography combined with flow injection chemiluminescence detection (HPLC-FI-CL) has been developed. The extracts were prepared by using methanol as extraction medium and two types of extraction methods (reflux and ultrasound assisted extraction). The post-column CL determination of polyphenols was based on their enhancing effect on the chemiluminescence intensity generated in manganese(IV)–hexametaphosphate–formaldehyde system in a phosphoric acid medium. Main antioxidants determined in C. palustre leaves were eriodictyol-7- O -glucoside, luteolin-7- O -glucoside and 6-hydroxyluteolin-7- O -glucoside belonging to flavonoids, and chlorogenic acid belonging to phenolic acids. Chromatographic separation was carried out on a C18 column with gradient elution by using a mobile phase containing 0.25% (v/v) phosphoric acid in water (solvent A) and 100% methanol (solvent B). Under the optimized conditions of chromatographic separation and CL detection the validation of the method was performed. The calibration curves showed good linearity in the concentration range from 0.5 to 40 µg mL −1 . The HPLC-FI-CL method was successfully applied to the determination of four polyphenolic compounds in methanolic extracts from leaves of C. palustre . The accuracy of the developed method was confirmed by the comparison of the results with those obtained by an HPLC-PDA method. The relative error of determination does not exceed 6.1%. However, the HPLC-FI-CL method is characterized by 40–65 times higher sensitivity compared to the HPLC-PDA method.

The determination of the sum of flavonoid compounds in extracts from inflorescences (expressed as mgL(-1) of apigenin) and leaves (expressed as mgL(-1) of linarin) of Cirsium oleraceum and Cirsium rivulare species by flow injection system with chemiluminescence detection (FI-CL) has been carried out. The method is based on the strong enhancement by polyphenols occurring in both plants of the CL signal generated by the reaction of cerium(IV) with rhodamine 6G in a sulfuric acid medium. Under the optimized conditions, the linear working ranges of 0.1-10 and 2.5-50μmolL(-1) were obtained for apigenin and linarin, respectively. The developed method is simple, sensitive with the detection limits of 38nmolL(-1) (apigenin) and 840nmolL(-1) (linarin) and offers high sample throughput (up to 300 samples per hour). The relative standard deviation was 0.62% and 3.75% for 10 measurements of 5μmolL(-1) apigenin and linarin, respectively. The proposed method has been successfully applied to determine the flavonoids/antioxidant levels in aqueous and methanolic extracts from inflorescences and leaves of C. oleraceum and C. rivulare. A possible mechanism of the enhancement of cerium(IV)-rhodamine 6G CL system by polyphenols was briefly discussed. For comparative studies, the antioxidant activity of C. oleraceum and C. rivulare extracts was also evaluated by spectrophotometric 1,1-diphenyl-2-picryl-hydrazyl (DPPH) radical scavenging method.

We recently reported that, in osteogenesis imperfecta (OI) type I with diminished type I collagen biosynthesis, flavonoids such as apigenin 7-O-glucuronide, apigenin 7-O-methylglucuronide and pectolinarin normalized the level of collagen type I without affecting total protein synthesis. In addition to collagen, glycosaminoglycans (GAGs) play an important role in the formation of a functional supramolecular complex in the extracellular matrix, and any changes in their content and/or composition may be involved in the OI phenotype. We previously detected a marked increase in sulphated GAG content in the OI fibroblasts of more severely affected patients (OI types II and III). These alterations were more pronounced in medium than in cells. Although, in OI type I cells, the increase observed in medium was much smaller (approximately 1.5-fold), it resulted in an increase of approximately 3-fold of the GAG to collagen type I ratio. Therefore, in the potential pharmacotherapy of OI type I with flavonoids, their effect on GAG level may be of importance. In the OI cells, some of the tested flavonoids applied at a concentration of 30 µM affected GAG content in quite the opposite way than type I collagen. Aglicones inhibiting collagen synthesis caused a marked increase in GAG concentration in medium, in contrast to the flavonoid glycosides, which exerted a stimulatory effect on type I collagen synthesis, but had a different effect on GAG content and distribution. Among these, apigenin 7-O-methylglucuronide did not affect GAG level or secretion, and thus may potentially be used in OI type I pharmacotherapy in patients with normal GAG content. However, in patients with increased concentrations of GAG, pectolinarin, which decreases GAG content by approximately 40%, may be more beneficial.

Aim of the study Cirsium rivulare (Jacq.) All. (Asteraceae) is a herbaceous perennial plant occurring in Central Europe. It has been traditionally used in Polish folk medicine to treat anxiety. In the present study methanolic extracts from flowers and leaves of Cirsium rivulare containing flavonoid compounds linarin, pectolinarin, apigenin, hispidulin, their glycosides and a newly isolated compound isokaemferide 7- O -(6″-methylglucuronide) were studied for anxiolytic and pro-cognitive properties. Materials and methods Male Wistar rats (150–160 g) were used. They were treated orally with standardized methanol extracts of flowers and leaves of Cirsium rivulare and subsequently tested for memory in passive avoidance (PA) and object recognition (OR) tests. Auxiliary tests for motor (open field, OF) and emotional (elevated ‘plus’ maze, EPM) effects of the above treatments were also employed. Results We found that the extract from flowers of Cirsium rivulare , in addition to its anxiolytic effects as measured in the EPM, improves memory of the appetitively (by curiosity, OR) and aversively (by footshook, PA) motivated tasks. This is in contrast to classical anxiolytics as for example benzodiazepines that typically impair memory. The extract from leaves of Cirsium rivulare showed some anxiolytic properties in the EPM, and no effect in both cognitive tests. The examined extracts of Cirsium rivulare did not affect psychomotor exploratory activity of rats tested in the OF. Conclusions These results suggest that the flavonoids from Cirsium rivulare possess anxiolytic and pro-cognitive effects, the extract from flowers being more pro-cognitive and that from the leaves more anxiolytic.

Crude aqueous extracts from leaves of Cirsium arvense, C. oleraceum, C. palustre, C. rivulare and C. vulgare were investigated. The content of tannins in mentioned sources, determined by the weight method with hide powder, varied between 1 and 7.63%. Total phenolic content, analysed by using Folin-Ciocalteau's method, ranged between 54 and 96 mg g(-1), was expressed as milligrams of gallic acid per gram of dry extract. Phenolic acids were identified by HPLC method. Antimicrobial activity of those extracts was examined. Cirsium palustre extract was the most active against investigated microorganisms. It was observed that the content of small-molecular phenolic compounds had greater influence on the activity of extracts than tannins. The total antioxidant activity indicated by radical cation 2,2'-azini-di-(3-ethylbenzthiazoline sulphonate) ABTS(*+), expressed as total antioxidant status (TAS) ranged from 2.31 to 2.78 mM L(-1).

It was found that the chemiluminescence (CL) produced from the reaction of luminol with iodine in the alkaline medium was strongly inhibited by plant phenolic compounds. Based on this finding, a new flow injection CL method was developed for the determination of caffeic acid and 6′-caffeoylerigeroside. The latter compound was isolated for the first time from Erigeron acris L. herb. The method was simple, rapid and sensitive with a detection limit of 4 × 10 −3 ng mL −1 (caffeic acid) and 0.18 ng mL −1 (6′-caffeoylerigeroside), linear range of 0.1–1.5 ng mL −1 (caffeic acid) and 1–200 ng mL −1 (6′-caffeoylerigeroside), relative standard deviation of 3.3% for 10 measurements of 0.45 ng mL −1 caffeic acid and 2.9% for 40 ng mL −1 6′-caffeoylerigeroside. This method was successfully applied to determine the content of phenolic compounds/antioxidant activity of E. acris L. extracts and phenolic acids content in pharmaceutical formulation. A possible mechanism of the inhibition of the proposed CL system was discussed.

The aim of the present study was to identify bioactive compounds stimulating collagen biosynthesis with potential for osteogenesis imperfecta (OI) type I pharmacological therapy. Of the compounds tested, apigenin glycosides 7-O-glucuronide, 7-O-methylglucuronide and pectolinarin at 30 microM were found to significantly induce collagen type I synthesis in OI fibroblasts without an effect on the overall protein synthesis. None of the compounds displayed any toxicity at that concentration. Secretion of collagen into media was not affected by apigenin 7-O-glucuronide and was slightly increased in cells treated with apigenin 7-O-methylglucuronide and pectolinarin. Furthermore, procollagen secreted by treated cells underwent a more rapid processing into collagen as compared with control untreated cells. In addition, we elucidated the possible mechanism involved in their action. Stimulation of collagen biosynthesis was not due to an increase in cell proliferation, because no differences in DNA content between the compound-treated and untreated cells were observed. Since flavonoids are known as strong inhibitors of metalloproteinases degrading matrix proteins, the increased level of collagen could result from the inhibition of their activity. However, the compounds with a stimulatory effect on collagen synthesis did not influence the activities of collagenase type I, gelatinases A and B, and stromelysin. On the contrary, all compounds stimulated the activity of prolidase which catalyzes the final step of collagen degradation and plays an important role in collagen biosynthesis. Stimulation of collagen synthesis and prolidase activity by apigenin 7-O-glucuronide was accompanied by an increase in IGF-I receptor expression. In contrast, the compounds apigenin 7-O-methylglucuronide and pectolinarin which normalized collagen synthesis in OI cells may exert their effects through beta1-integrin-mediated signaling.

The essential oil from Erigeron acris L. obtained by hydrodistillation was analyzed by GC and GC/MS techniques. Over 60 compounds were identified, representing 92.6% of the total components. The major constituents of the oil were monoterpenoid hydrocarbons [limonene, β-pinene, (E)-β-ocimene] and sesquiterpenoid hydrocarbons [α-muurolen, germacrene D, (E)-β-farnesene].

The composition of the essential oils of Cirsium palustre and C. rivulare and their antiproliferative activity against breast adenocarcinoma cells (MCF-7 and MDA-MBA-231) were investigated. The essential oils obtained by hydro-distillation from the roots (yield 0.2 and 0.1% v/w, respectively), leaves and inflorescences (yield below 0.01%), were analyzed by gas chromatography coupled with mass spectrometry (GC-MS). The composition of the essential oils in the respective organs of each plant differed considerably. On the other hand, similarities were observed in the composition of root and leaf oils. In C. palustre and C. rivulare root oil, 50 and 39 constituents were identified, accounting for 95.3% and 92.4% of the total content. The main components were aplotaxene and its derivatives, representing 78.6% and 46.6% of the total content. In leaf oils of both species, 59 and 49 compounds, respectively, were identified, representing 67.4% and 78.3% of the total content. The major constituents were β-damascenone (4.1% and 13.4%, respectively) and β-ionone (6.7% and 5.3%, respectively). Short-chain saturated and unsaturated aliphatic alcohols and aldehydes constituted another important group of compounds (17.7% and 9.0%, respectively). The essential oils of the roots have moderate anti-proliferative activity, with IC50 values ranging from 110 to 140 μg/mL. These concentrations were below the level able to inhibit the proliferation of healthy cells.

The collagen metabolism alterations triggered by reactive oxygen species are involved in the development of various connective tissue diseases and skin aging. This study was designed to examine whether (E)-anethole possesses a protective effect on H2O2-induced alterations in collagen metabolism as well as whether it can prevent apoptosis in human skin fibroblasts. In cells treated with 300 µM H2O2, a decrease in collagen biosynthesis of 54 % was observed. Pretreatment of cells with 0.5 µM anethole for 1 h completely prevented this alteration. Changes at the protein level positively correlated with alterations of type I collagen mRNA expression. We have shown that H2O2 caused increase in the activity of MMP-2 and MMP-9 as well as that an increase in MMP-2 activity can contribute to the 8 % decrease in the amount of collagen secreted into the medium. The most efficient suppression of these changes was observed in the presence of 0.5 µM of anethole. At 10 µM, in addition to suppression, an inhibitory effect of anethole on MMP-9 activity was documented. Additionally, the 60 % H2O2-induced decrease in cell viability was suppressed by 1 µM of anethole and a 4-fold increase in cell apoptosis was suppressed by 0.5 µM of anethole. Our results suggest that anethole, which is a small lipophilic and non-toxic molecule with the ability to prevent H2O2-induced collagen metabolism alterations and apoptosis in human skin fibroblasts, would prove useful in the development of effective agents in pharmacotherapy of oxidative stress-related skin diseases.

The chemical composition of Cirsium vulgare flower heads was examined. Petrol and chloroform extracts of this plant material were analyzed by GC-MS for the presence of fatty acids, phytosterols, and triterpenes. Diethyl ether, ethyl acetate, and butanol fractions of the methanolic extract were subjected to multistep chromatographic separations, and as a result, fourteen flavonoids were obtained (1–14). All compounds were isolated from this morphological part for the first time and eleven from the plant. Among the identified components were four aglycones, eight glycosides, and two glycoside esters, derivatives of apigenin, luteolin, kaemferol, and quercetin. One of them was a rarely occurring compound apigenin 7- O-β-(6″-butyl)-glucuronide) (14). Total flavonoid content and antioxidant activity were determined in the various fractions of methanolic extract.