Determination of Selected Isoquinoline Alkaloids from Chelidonium majus, Mahonia aquifolium and Sanguinaria canadensis Extracts by Liquid Chromatography and Their In Vitro and In Vivo Cytotoxic Activity against Human Cancer Cells.

The search for new substances with cytotoxic activity against various cancer cells, especially cells that are very resistant to currently used chemotherapeutic agents, such as melanoma cells, is a very important scientific aspect. We investigated the cytotoxic effect of Chelidonium majus, Mahonia aquifolium and Sanguinaria canadensis extracts obtained from different parts of these plants collected at various vegetation stages on FaDu, SCC-25, MCF-7, and MDA-MB-231 cancer cells. Almost all the tested extracts showed higher cytotoxicity against these cancer cells than the anticancer drug etoposide. The highest cytotoxicity against the FaDu, SCC-25, MCF-7 and MDA-MB-231 cancer cell lines was obtained for the Sanguinaria candensis extract collected before flowering. The cytotoxicity of extracts obtained from different parts of Chelidonium majus collected at various vegetation stages was also evaluated on melanoma cells (A375, G361 and SK-MEL-3). The highest cytotoxic activity against melanoma A375 cells was observed for the Chelidonium majus root extract, with an IC50 of 12.65 µg/mL. The same extract was the most cytotoxic against SK-MEL-3 cells (IC50 = 1.93 µg/mL), while the highest cytotoxic activity against G361 cells was observed after exposure to the extract obtained from the herb of the plant. The cytotoxic activity of Chelidonium majus extracts against melanoma cells was compared with the cytotoxicity of the following anticancer drugs: etoposide, cisplatin and hydroxyurea. In most cases, the IC50 values obtained for the anticancer drugs were higher than those obtained for the Chelidonium majus extracts. The most cytotoxic extract obtained from the root of Chelidonium majus was selected for in vivo cytotoxic activity investigations using a Danio rerio larvae xenograft model. The model was applied for the first time in the in vivo investigations of the extract's anticancer potential. The application of Danio rerio larvae xenografts in cancer research is advantageous because of the transparency and ease of compound administration, the small size and the short duration and low cost of the experiments. The results obtained in the xenograft model confirmed the great effect of the investigated extract on the number of cancer cells in a living organism. Our investigations show that the investigated plant extracts exhibit very high cytotoxic activity and can be recommended for further experiments in order to additionally confirm their potential use in the treatment of various human cancers.

Polyphenol Composition and Antioxidant Potential of Instant Gruels Enriched with Lycium barbarum L. Fruit.

Goji fruit (Lycium barbarum L.) has been identified as a polyphenolic compound plant source of noted richness. It also contains polysaccharides, carotenoids, vitamins and minerals, fatty and organic acids. The purpose of the presented research was to produce innovative instant corn gruels with various dry goji berry contents (1, 3 and 5%), to determine the level of included polyphenolic compounds (including individual free phenolic acids) and to assess the antioxidant properties of these functional-food products. A further objective was to identify the optimum value of one of the most important production parameter, the rotational speed of the extruder's screw during gruel processing. The undertaken chromatographic analysis (LC-ESI-MS/MS) showed a wide variety of available phenolic acids. In the samples with 5% addition of fruit, eight phenolic acids were detected, whereas in the corn gruel without additives, only five were noted. The antioxidant activity, the content of free phenolic acids and the sum of polyphenols increased with increase of the functional additive. For all goji content, screw speeds of 100 and 120 rpm rather than 80 rpm resulted in higher polyphenol amounts and greater Trolox equivalent antioxidant capacity, as well as higher ability to scavenge DPPH.

ANALYSIS OF BASIC PSYCHOTROPIC DRUGS IN BIOLOGICAL FLUIDS AND TISSUES BY REVERSED-PHASE HIGH PERFORMANCE LIQUID CHROMATOGRAPHY.

The review of the RP HPLC analysis of basic psychotropic drugs is presented. It contains sample preparation methods with centrifugation, protein precipitation, liquid-liquid extraction (LLE), dispersive liquid-liquid microextraction (DLLME), solid-phase extraction (SPE), solid-phase microextraction (SPME), microwave-assisted extraction (MAE) and RP-HPLC analysis. Chromatographic behavior of basic drugs in aqueous media - eluents used in reversed phase systems is discussed. Methods of blocking of residue surface silanols' interaction are mentioned. Analytical methods used for the analysis are divided into parts according with the above methods: the use of low-pH eluents, the use of high-pH eluents, the use of silanol blockers, special stationary phases for basic analytes. Literature connected with the sample preparation methods and analytical systems for the drug analysis are cited in details and presented also in Table 1.

RP-HPLC ANALYSIS OF ACIDIC AND BASIC DRUGS IN SYSTEMS WITH DIETHYLAMINE AS ELUENTS ADDITIVE.

The chromatographic behavior of some basic and acidic drugs was studied on Cl 8, Phenyl-Hexyl and Polar RP columns with methanol or acetonitrile as organic modifiers of aqueous mobile phases containing addition of diethylamine. Diethylamine plays a double function of silanol blocker reagent in analysis of basic drugs and ion-pair reagent in analysis of acidic drugs. Most symmetrical peaks and highest system efficiency were obtained on Phenyl-Hexyl and Polar RP columns in tested mobile phase systems compared to results obtained on C18 column. A new rapid, simple, specific and accurate reverse phase liquid chromatographic method was developed for the simultaneous determination of atorvastatin - antihyperlipidemic drug and amlodipine - calcium channel blocker in one pharmaceutical formulation. Atorvastatin is an acidic compounds while amlodipine is a basic substance. The chromatographic separation was carried out on Phenyl-Hexyl column by gradient elution mode with acetonitrile as organic modifier, acetate buffer at pH 3.5 and Q.025 M/L diethylamine. The proposed method was validated for specificity, precision, accuracy, linearity, and robustness. The linearity range of atorvastatin and amlodipine for 5 - 100 µg/mL was obtained with limits of-detection (LOD) 3.2750 gg/mL and 3.2102 µg/mL, respectively. The proposed method made use of DAD as a tool for peak identity and purity confirmation.

Ion-exchange vs reversed-phase chromatography for separation and determination of basic psychotropic drugs.

Ion exchange chromatography, an alternative to reversed-phase (RP) chromatography, is described in this paper. We aimed to obtain optimal conditions for the separation of basic drugs because silica-based RP stationary phases show silanol effect and make the analysis of basic analytes hardly possible. The retention, separation selectivity, symmetry of peaks and system efficiency were examined in different eluent systems containing different types of buffers at acidic pH and with the addition of organic modifiers: methanol and acetonitrile. The obtained results reveal a large influence of the salt cation used for buffer preparation and the type of organic modifier on the retention behavior of the analytes. These results were also compared with those obtained on an XBridge C18 column. The obtained results demonstrated that SCX stationary phases can be successfully used as alternatives to C18 stationary phases in the separation of basic compounds. The most selective and efficient chromatographic systems were applied for the quantification of some psychotropic drugs in fortified human serum samples.

Antioxidant Activity of Selected Thyme (Thymus L.) Species and Study of the Equivalence of Different Measuring Methodologies.

This study presents the results of comparative evaluation of the antioxidant activity of the phenolic fraction exhaustively extracted with aqueous methanol from 18 different thyme (Thymus L.) specimens and species. This evaluation is made with use of the same free radical source (DPPH• radical), three different free radical scavenging models (gallic acid, ascorbic acid, and Trolox), and three different measuring techniques (the dot blot test, UV-Vis spectrophotometry, and electron paramagnetic resonance spectroscopy, EPR). A comparison of the equivalence of these three different measuring techniques (performed with use of hierarchical clustering with Euclidean distance as a similarity measure and Ward's linkage) is particularly important in view of the fact that different laboratories use different antioxidant activity measuring techniques, which makes any interlaboratory comparison hardly possible. The results obtained confirm a semiquantitative equivalence among the three compared methodologies, and a proposal is made of a simple and cost-effective dot blot test that uses the DPPH• radical and provides differentiation of antioxidant activity of herbal matter comparable with the results of the UV-Vis spectrophotometry and EPR.

A Comparison of Antibacterial Activity of Selected Thyme (Thymus) Species by Means of the Dot Blot Test with Direct Bioautographic Detection.

Bioautography carried out with the aid of thin-layer chromatographic adsorbents can be used to assess antibacterial activity in samples of different origin. It can either be used as a simple and cost-effective detection method applied to a developed chromatogram, or to the dot blot test performed on a chromatographic plate, where total antibacterial activity of a sample is scrutinized. It was an aim of this study to compare antibacterial activity of 18 thyme (Thymus) specimens and species (originating from the same gardening plot and harvested in the same period of time) by means of a dot blot test with direct bioautography. A two-step extraction of herbal material was applied, and at step two the polar fraction of secondary metabolites was obtained under the earlier optimized extraction conditions [methanol-water (27+73, v/v), 130°C]. This fraction was then tested for its antibacterial activity against Bacillus subtilis bacteria. It was established that all investigated extracts exhibited antibacterial activity, yet distinct differences were perceived in the size of the bacterial growth inhibition zones among the compared thyme species. Based on the results obtained, T. citriodorus "golden dwarf" (sample No. 5) and T. marschallianus (sample No. 6) were selected as promising targets for further investigations and possible inclusion in a herbal pharmacopeia, which is an essential scientific novelty of this study.

Effect of chromatographic conditions on retention behavior and system efficiency for HPTLC of selected psychotropic drugs on chemically bonded stationary phases.

Selected psychotropic drug standards have been chromatographed on RP18, CN and diol layers with a variety of aqueous and nonaqueous mobile phases. The effect of buffers at acidic or basic pH, acetic acid, ammonia and diethylamine (DEA) in aqueous mobile phases on retention, efficiency and peak symmetry was examined. Improved peak symmetry and separation selectivity for investigated compounds were observed when ammonia or DEA were used as mobile phase additives. The effect of diethylamine concentration in aqueous eluents on retention, peak symmetry and theoretical plate number obtained on CN plates was also investigated. Because of the strong retention of these basic drugs on stationary phases bonded on silica matrix, nonaqueous eluents containing medium polar diluents, strongly polar modifiers and silanol blockers (ammonia or diethylamine) were applied. Aqueous and nonaqueous eluent systems with the best selectivity and efficiency were used for separate psychotropic drug standards' mixture on CN layer by 2D TLC.

The use of TLC-DPPH* test with image processing to study direct antioxidant activity of phenolic acid fractions of selected Lamiaceae family species.

TLC coupled with 2,2-diphenyl-1-picrylhydrazyl staining was used to analyze phenolic acid fractions of selected Salvia and Thymus species. Documented videoscans were processed by means of an image processing program. This is the first time that free phenolic acids fractions, as well as fractions containing phenolic acids derived from basic and acidic hydrolysis, have been analyzed and compared for selected sage and thyme species. The analyzed samples along with caffeic acid (CA; standard) were chromatographed on silica gel plates with toluene-ethyl acetate-formic acid (60 + 40 + 1, v/v/v) mobile phase. The extracts were investigated with respect to the activity of CA. It was found that CA was most abundant in the fractions derived from basic hydrolysis. This compound was not detected in any of the fractions obtained after acidic hydrolysis. S. officinalis and S. triloba have similar free radical scavenging activity fingerprints obtained for all the analyzed fractions.

Micro 2D-TLC of Selected Plant Extracts in Screening of Their Composition and Antioxidative Properties.

Micro two-dimensional separations were performed on polar bonded stationary phases of the type cyanopropyl-silica using non-aqueous eluents (polar modifier dissolved in n-heptane) as the first direction eluents and aqueous eluents (organic modifier-MeOH dissolved in water) as the second direction eluents. The chromatographic process was performed in micro scale using 5 × 5 cm plates, small volumes of eluents and 10 µL of plant extracts to obtain satisfying separation. Plates developed in horizontal chambers were dried and observed in UV light (254 nm and 366 m) photographed by digital camera and derivatized by DPPH to detect antioxidants (free radical scavengers) or derivatized by Naturstoff reagent to detect phenolic compounds (characteristic luminescence of some phenolic compounds). The above experiments give the possibility to construct fingerprints for investigated Polygonum hydropiper, Betula verrucosa and Pulmonaria officinalis extracts. It can be used in quality control of the plant material and its antioxidative activity. Novelty of the paper is the micro-scale of the separation by two-dimensional thin layer chromatography mode. For the first time two-dimensional separation of plant extracts on 5 × 5 cm plates in two directions is performed.

Binary HPLC-diode array detector and HPLC-evaporative light-scattering detector fingerprints of methanol extracts from the selected sage (Salvia) species.

This study is focused on an important family of the sage (Salvia) species, with Salvia officinalis L. having a long-established position in European traditional medicine. Binary fingerprints (chromatographic profiles) of six different sage species were compared using HPLC coupled with two different detectors: the diode-array detector and the evaporative light-scattering detector. Advantages of using binary fingerprinting over single-detector fingerprinting are demonstrated and discussed, with selected examples. Experimental data are provided for a comparison of the chemical composition of sage samples originating from two harvesting seasons (2007 and 2008). A number of phytochemical standards (i.e., certain phenolic acids, flavonoids, and coumarin) were used that allowed identification and semiquantitative estimation of these particular compounds in the analyzed methanol extracts.

Development of chromatographic and free radical scavenging activity fingerprints by thin-layer chromatography for selected Salvia species.

INTRODUCTION: Plant-derived free radical scavengers have become the subject of intensive scientific interest. Recently, the concept of coupling chromatographic fingerprints with biological fingerprinting analysis has gained much attention for the quality control of plant extracts. However, identification of free radical scavenging activity of each single compound in a complex mixture is a difficult task. Thin-layer chromatography with post-chromatographic derivatisation with the methanol solution of DPPH can be a valuable tool in such analyses. OBJECTIVE: Development of chromatographic and free radical scavenging fingerprints of nineteen Salvia species grown and cultivated in Poland. METHODOLOGY: Chromatography was performed on the silica gel layers with use of two eluents, one for the resolution of the less polar compounds, and the other one for the resolution of the medium and highly polar ones. The plates were sprayed with the vanillin-sulfuric acid reagent to produce chemical fingerprints, and with DPPH solution to generate free radical scavenging fingerprints. RESULTS: With four Salvia species, it was revealed that their strong free radical scavenging properties are not only due to the presence of polar flavonoids and phenolic acids, but also due to the presence of several free radical scavengers in the less polar fraction. Because of the similarities in both the chromatographic and the free radical scavenging fingerprints, S. triloba can be introduced as a possible equivalent of the pharmacopoeial species, S. officinalis. CONCLUSION: Fingerprints developed in the experiments proved useful for the analysis of complex extracts of the different Salvia species.

Application of thin-layer chromatography for the quality control and screening the free radical scavenging activity of selected pharmacuetical preparations containing Salvia officinalis L. extract.

Thin-layer chromatographic method, with postchromatographic derivatization, was applied for the purposes of the quality control of pharmaceutical preparations, containing S. officinalis L. extract. Six finished products underwent the analysis: capsules, tablets, two ointments, tincture and finished product being a mixture of ethanolic S. officinalis and Thymi vulgaris extracts. Chromatographic and free radical scavenging fingerprints, obtained for the herbal products, were compared with the profiles of the authenticated botanical reference material. The application of the proposed technique revealed most of the fingerprints, developed for the analyzed preparations, matched with the profiles obtained for authenticated plant material. The developed method was found suitable for the quality control of herbal preparations containing sage extract.

Determination of Cytisine and N-Methylcytisine from Selected Plant Extracts by High-Performance Liquid Chromatography and Comparison of Their Cytotoxic Activity.

Quinolizidine alkaloids exhibit various forms of biological activity. A lot of them were found in the Leguminosae family, including Laburnum and Genista. The aim of the study was the optimization of a chromatographic system for the analysis of cytisine and N-methylcytisine in various plant extracts as well as an investigation of the cytotoxic activities of selected alkaloids and plant extracts obtained from Laburnum anagyroides, Laburnum anagyroides L. quercifolium, Laburnum alpinum, Laburnum watereri, Genista germanica, and Genista tinctoria against various cancer cell lines. The determination of investigated compounds was performed by High Performance Liquid Chromatography with Diode Array Detection (HPLC-DAD), while High Performance Liquid Chromatography coupled with Quadrupole Time-of-Flight-Mass Spectrometry (HPLC-QTOF-MS) was applied for the qualitative analysis of plant extracts. The retention, separation selectivity, peaks shape, and systems efficiency obtained for cytisine and N-methylcytisine in different chromatographic systems were compared. The application of columns with alkylbonded and phenyl stationary phases led to a very weak retention of cytisine and N-methylcytisine, even when the mobile phases containing only 5% of organic modifiers were used. The strongest retention was observed when hydrophilic interaction chromatography (HILIC) or especially when ion exchange chromatography (IEC) were applied. The most optimal system in terms of alkaloid retention, peak shape, and system efficiency containing an strong cation exchange (SCX) stationary phase and a mobile phase consisted of 25% acetonitrile and formic buffer at pH 4.0 was applied for investigating alkaloids analysis in plant extracts. Cytotoxic properties of the investigated plant extracts as well as cytisine and N-methylcytisine were examined using human tongue squamous carcinoma cells (SCC-25), human pharyngeal squamous carcinoma cells (FaDu), human triple-negative breast adenocarcinoma cell line (MDA-MB-231), and human breast adenocarcinoma cell line (MCF-7). The highest cytotoxic activity against FaDu, MCF-7, and MDA-MB cancer cell lines was observed after applying the Genista germanica leaves extract. In contrast, the highest cytotoxic activity against SCC-25 cell line was obtained after treating with the seed extract of Laburnum watereri. The investigated plant extracts exhibit significant cytotoxicity against the tested human cancer cell lines and seem to be promising for further research on its anticancer activity.

Two-dimensional thin-layer chromatography in the analysis of secondary plant metabolites.

Drugs, derived from medicinal plants, have been enjoying a renaissance in the last years. It is due to a great pharmacological potential of herbal drugs, as many natural compounds have been found to exhibit biological activity of wide spectrum. The introduction of whole plants, plant extracts, or isolated natural compounds has led to the need to create the analytical methods suitable for their analysis. The identification of isolated substances is relatively an easy task, but the analysis of plant extracts causes a lot of problems, as they are usually very complex mixtures. Chromatographic methods are one of the most popular techniques applied in the analysis of natural mixtures. Unfortunately the separation power of traditional, one-dimensional techniques, is usually inadequate for separation of more complex samples. In such a case the use of multidimensional chromatography is advised. Planar chromatography gives the possibility of performing two-dimensional separations with the use of one adsorbent with two different eluents or by using bilayer plates or graft thin-layer chromatography (TLC) technique; combinations of different multidimensional techniques are also possible. In this paper, multidimensional planar chromatographic methods, commonly applied in the analysis of natural compounds, were reviewed. A detailed information is given on the methodology of performing two-dimensional separations on one adsorbent, on bilayer plates, with the use of graft TLC and hyphenated methods. General aspects of multidimensionality in liquid chromatography are also described. Finally a reader will find a description of variable two-dimensional methods applied in the analysis of compounds, most commonly encountered in plant extracts. This paper is aimed to draw attention to the potential of two-dimensional planar chromatography in the field of phytochemistry. It may be useful for those who are interested in achieving successful separations of multicomponent mixtures by means of two-dimensional TLC.

Fingerprint of selected Salvia species by HS-GC-MS analysis of their volatile fraction.

Twenty species of Salvia, naturally grown or cultivated in Poland, are investigated by headspace gas chromatography-mass spectrometry analysis. The main components of the volatile fraction of Salvia species are identified as alpha-pinene, camphene, beta-pinene, thujol, camphor, beta-chamigrene, and cadina-3,9-diene. There are also the compounds that can be considered as chemotaxonomic markers, namely beta-myrcene for Salvia lavadulifolia, beta-phelandrene for Salvia verticillata, tau-terpinene for Salvia stepposa, and isocaryophyllene and caryophyllene for Salvia officinalis. Certain compounds (such as o-cymene present in Salvia canariensis and Salvia stepposa; beta-trans-ocymene present in Salvia lavadulifolia, Salvia sclarea, and Salvia amplexicaulis; thujenone present in Salvia staminea, Salvia atropatana, Salvia jurisicii, and Salvia officinalis; and thujone present in Salvia azurea, Salvia lavandulifolia, Salvia hians, and Salvia triloba) can constitute chemotaxonomic advice for the aforementioned species. Also, the lack of certain compounds otherwise common in the individual sage species can be considered as chemotaxonomic advice (e.g., Salvia sclarea has no alpha-pinene and beta-pinene; Salvia lavadulifolia lacks camphene; Salvia triloba lacks beta-pinene and camphene; and Salvia officinalis lacks beta-chamigrene, thujol, and cadina-3,9-diene).

Liquid chromatography fingerprint analysis and antioxidant activity of selected lavender species with chemometric calculations.

The extracts of seven Lavandulae species (Lavandula stoechas, Lavandula lanata, Lavandula viridis, Lavandula angustifolia "Rosea", Lavandula angustifolia "Afropurpurea", Lavandula angustifolia and one unknown) were analyzed using the reversed-phase-high performance liquid chromatography-diode array detection (RP-HPLC-DAD) with gradient elution technique to obtain the chromatographic fingerprint profiles. The HPLC analysis was performed using the Kinetex RP18 chromatographic column and eluent consisting of methanol-water-0.1% formic acid (5-100% (v/v)) at 30 °C with the run time of 60 min. and the detection wavelength 280 nm. The chromatograms were preliminary processed with the smoothing, noise reduction, background subtraction and alignment using the SpecAlign program (version 2.4.1). The presence of selected standards (apigenin, myricetin, luteolin, luteolin 7-glucoside, chlorogenic acid, caffeic acid, ferulic acid) in the extracts was confirmed. The chemical similarity between studied plants was evaluated using the Cluster Analysis (Pearson correlation coefficient, r, and Euclidean) and PCA. The preliminary antioxidant activity of studied extracts was evaluated based on the total phenolic content (Folin-Ciocalteu method), ferric ion reducing antioxidant parameter (FRAP) and α,α-diphenyl-ß-picrylhydrazyl (DPPH) free radical scavenging method using the spectrophotometric technique.

An attempt to elucidate the role of iron and zinc ions in development of Alzheimer's and Parkinson's diseases.

Neurodegenerative disorders are among the most studied issues both in medicine and pharmacy. Despite long and extensive research, there is no effective treatment prescribed for such diseases, including Alzheimer's or Parkinson's. Available data exposes their multi-faceted character that requires a complex and multidirectional approach to treatment. In this case, the most important challenge is to understand the neurodegenerative mechanisms, which should permit the development of more elaborate and effective therapies. In the submitted review, iron and zinc are discussed as important and perfectly possible neurodegenerative factors behind Alzheimer's and Parkinson's diseases. It is commonly known that these elements are present in living organisms and are essential for the proper operation of the body. Still, their influence is positive only when their proper balance is maintained. Otherwise, when any imbalance occurs, this can eventuate in numerous disturbances, among them oxidative stress, accumulation of amyloid ß and the formation of neurofibrillary tangles, let alone the increase in α-synuclein concentration. At the same time, available research data reveals certain discrepancies in approaching metal ions as either impassive, helpful, or negative factors influencing the development of neurodegenerative changes. This review outlines selected neurodegenerative disorders, highlights the role of iron and zinc in the human body and discusses cases of their imbalance leading to neurodegenerative changes as shown in vitro and in vivo studies as well as through relevant mechanisms.

Determination of Selected Isoquinoline Alkaloids from Mahonia Aquifolia; Meconopsis Cambrica; Corydalis Lutea; Dicentra Spectabilis; Fumaria Officinalis; Macleaya Cordata Extracts by HPLC-DAD and Comparison of Their Cytotoxic Activity.

Alkaloids have protective functions for plants and can play an important role in living organisms. Alkaloids may have a wide range of pharmacological activities. Many of them have cytotoxic activity. Nowadays, cancer has become a serious public health problem. Searching for effective drugs with anticancer activity is one of the most significant challenges of modern scientific research. The aim of this study was the investigation of cytotoxic activity of extracts obtained from Corydalis lutea root and herb, Dicentra spectabilis root and herb, Fumaria officinalis, Macleaya cordata leaves and herb, Mahonia aquifolia leaves and cortex, Meconopsis cambrica root and herb on FaDu, SCC-25, MCF-7, and MDA-MB-231 cancer cell lines. The cytotoxic activity of these extracts has not been previously tested for these cell lines. The aim was also to quantify selected alkaloids in the investigated extracts by High Performance Liquid Chromatography (HPLC). The analyses of alkaloid content were performed using HPLC in reversed phase (RP) mode using Polar RP column and mobile phase containing acetonitrile, water and ionic liquid (IL). Cytotoxic effect of the tested plant extracts and respective alkaloid standards were examined using human pharyngeal squamous carcinoma cells (FaDu), human tongue squamous carcinoma cells (SCC-25), human breast adenocarcinoma cell line (MCF-7), human triple-negative breast adenocarcinoma cell line (MDA-MB-231). All investigated plant extracts possess cytotoxic activity against tested cancer cell lines: FaDu, SCC-25, MCF-7, and MDA-MB-231. The highest cytotoxic activity against FaDu, SCC-25, and MCF-7 cell lines was estimated for Macleaya cordata leaf extract, while the highest cytotoxic activity against MDA-MB-231 cell line was obtained for Macleaya cordata herb extract. Differences in cytotoxic activity were observed for extracts obtained from various parts of investigated plants. In almost all cases the cytotoxic activity of investigated plant extracts, especially at the highest concentration against tested cell lines was significantly higher than the activity of anticancer drug etoposide. Our investigations exhibit that these plant extracts can be recommended for further in vivo experiments to confirm their anticancer activity.

Two-dimensional thin-layer chromatography with adsorbent gradient as a method of chromatographic fingerprinting of furanocoumarins for distinguishing selected varieties and forms of Heracleum spp.

There are a lot of taxonomic classifications of the genus Heracleum, and many authors indicate they need revision. Morphological identification is difficult to perform, as there are only few characteristic differences between each Heracleum species, varieties and forms. Furanocoumarins are characteristic compounds for the Apiaceae family, and they can be found in the whole genus in large quantities. Despite this fact, it is difficult to use the furanocoumarin profiles of plants, for their discrimination, as furanocoumarins are difficult to separate, due to their similar chemical structures and physicochemical properties. In this paper, a new, simple method is proposed for the discrimination of selected species, varieties and forms of the genus Heracleum. Thin-layer chromatography (TLC) with an adsorbent gradient (unmodified silica gel+octadecylsilica wettable with water) enables complete separation of the structural analogues. The proposed method gives the possibility to distinguish selected species, varieties and forms of the Heracleum genus, as they produce distinctive furanocoumarin fingerprints. The method is characterised by high specificity, precision, reproducibility and stability values. It is for the first time that graft TLC is used for constructing fingerprints of herbs. The complete separation of ten structural analogues, by combining gradient TLC with the unidimensional multiple development technique, has not been reported yet.