Taraxacum officinale extract shows antitumor effects on pediatric cancer cells and enhance mistletoe therapy.

OBJECTIVES: While Taraxacum officinale (dandelion) extracts showed antitumor effects on adult cancer cells, effects on pediatric tumor cells as a single agent or in combination with mistletoe extracts are hitherto unknown. MATERIAL AND METHODS: The anti-proliferative effects of an aqueous fermented Taraxacum officinale extract (Taraxacum) on a pediatric cancer cell line panel were assessed by cell viability assays (MTT). In two neuroblastoma cell lines, SH-SY5Y and Kelly, the effects on cell cycle distribution (PI staining), mitochondrial integrity (MitoTracker staining), invasion (Boyden chamber assay) and migration (Scratch-assay) as well as the synergistic effects of the co-treatment of Taraxacum and mistletoe preparations (Iscucin® Tiliae or Iscucin® Pini) were investigated. RESULTS: All tested cancer cell lines were more susceptible to Taraxacum than the normal human fibroblast cell line, NHDF-C. In neuroblastoma cell lines Taraxacum caused apoptosis and loss of mitochondrial integrity as well as an inhibition of invasion and migration. The simultaneous therapy of Taraxacum and the mistletoe extracts revealed synergistic effects. CONCLUSION: This preclinical data support the use of Taraxacum as a potential adjuvant application in pediatric oncology.

SOD activity and extremophilicity: a screening of various plant species.

All aerobes are dependent on enzymatic and non-enzymatic antioxidants to withstand the presence of reactive oxygen species (ROS). Superoxide dismutase (SOD) is a part of the enzymatic antioxidant system. It is one of the most important antioxidant enzymes, enabling organisms to survive in an oxygen containing atmosphere. A disorder in the oxidative and antioxidative balance can be associated with the occurrence of diseases in human organisms. Little data exist on the relevance of SOD in plants. Moreover, it is not known whether there is any association between a plant's origin and its SOD activity. Our screening of 27 different plant species was intended to expose whether there is a connection. The highest SOD activities were found for extremophile plants. Especially the Crassulaceae Aeonium haworthii Salm-Dyck Ex Webb & Berthel. and Crassula multiflora Schönland & Baker F. were highly active. Nevertheless, we did not find unambiguous evidence for a correlation between extremophilicity and SOD activity.

Comprehensive phytochemical characterization of St. John's wort (Hypericum perforatum L.) oil macerates obtained by different extraction protocols via analytical tools applicable in routine control.

The aim of the present study was to investigate the impact of crucial process parameters, i.e. of light and temperature conditions, during the preparation of St. John's wort (SJW, Hypericum perforatum L.) Arachis oil macerates. Extracts were prepared according to a standardized protocol over a period of 28 days. For this purpose, flowering tops of H. perforatum were macerated with Arachis oil (drug extract ratio, DERnative 1:4) under different light and temperature conditions. Spectrophotometric measurements were carried out to quantitate naphthodianthrones and to characterize extract color in the CIE L*C*h° system. Moreover, individual plant secondary metabolites were screened by UHPLC-DAD-MSn measurements following liquid-liquid extraction of the oil macerates with methanol. For quantitation purposes, the chromatographic method was validated using reference standards. This methodology allowed the separation of up to 25 constituents in oily and methanolic SJW extracts, covering hydroxycinnamic acids, flavanols, proanthocyanidins, flavonol glycosides, flavonol aglyca, biflavones, bisanthraquinone glycosides, naphthodianthrones and phloroglucinols. Lowest naphthodianthrone contents were determined in oil macerates recovered at 5 °C, whereas highest amounts were detected upon extraction at 50 °C (both under the exclusion of light). Color shades of the oil macerates differed markedly, revealing e. g. a*-values ranging from -4.6±0.3 to 42.5±0.3. The flavonoids quercetin, kaempferol and I3, II8-biapigenin as well as the phloroglucinols hyperforin and adhyperforin could be simultaneously detected and quantitated in all oil macerates. Contents of these constituents varied noticeably between macerates prepared under different conditions (quercetin 14.7±1.2 to 21.8±0.6 µg/g, kaempferol 3.0±0.1 to 5.4±0.4 µg/g, I3, II8-biapigenin 4.4±0.2 to 7.4±0.4 µg/g, hyperforin 52.6±46.0 to 451.4±24.9 µg/g, adhyperforin 6.9±5.7 to 74.5±7.1 µg/g). These results confirm that the quality of the resulting plant extracts is largely determined by the respective process parameters, i.e. especially temperature and light conditions, and thus must be thoroughly chosen and monitored to obtain tailor-made preparations.

Biochemical, microbiological and phytochemical studies on aqueous-fermented extracts from Atropa belladonna L. Part 2--Phytochemistry.

Extraction methods of fresh plants into aqueous-fermented extracts according to German Homoeopathic Pharmacopoeia (HAB), regulation nos. 33 and 34 were evaluated. In the course of production, the extraction is accompanied by fermentation and the resulting preparation is stored for at least 6 months until further processing. In part 1 of the work, the biochemical reactions proceeding in aqueous-fermented extracts from the fresh flowering herb of Atropa belladonna var. belladonna (L.) were studied and the responsible microorganisms were identified. This second part aimed at shedding light on the phytochemical changes upon manufacture and storage. Additionally, questions were addressed at the robustness of the method by varying production conditions, its reproducibility and the comparison with ethanolic extraction. Studying 110 extracts produced from 2006 to 2009 as well as model experiments on isolated lactic acid bacteria in atropine solutions proved that the active substance atropine was stable under regular pH conditions. Interestingly, no difference between aqueous-fermented and ethanolic extracts could be found with respect to atropine concentration. In contrast, the amounts of scopoletin and kaempferol glycosides from Atropa belladonna differed depending on the extraction procedure.

Time-dependent bioactivity of preparations from cactus pear (Opuntia ficus indica) and ice plant (Mesembryanthemum crystallinum) on human skin fibroblasts and keratinocytes.

ETHNOPHARMACOLOGICAL RELEVANCE: Traditionally and nowadays preparations from two xerophytic plants, the ice plant and cactus pear are used in dermatologic and cosmetic preparations. In spite of their daily use, little is known concerning the bioactivity of such extracts on skin cells. The purpose of this study was to investigate the effect of pressed juices from ice plant (McP) and two cactus pear polysaccharides (cold water soluble, NwPS; non swelling pectin, NPec) on the cell physiology of normal human dermal fibroblasts (NHDF) and HaCaT-keratinocytes due to composition, concentration and incubation time. MATERIALS AND METHODS: Cactus pear polysaccharides were analyzed by high performance anion exchange chromatography with pulsed amperometric detection after hydrolysis with trifluoroacetic acid. Ice plant pressed juices were filtrated through a 1.2 µm (McPI) and 0.2 µm filter (McPII). Cell proliferation was measured with BrdU incorporation assay. Reduction of tetrazolium salts was applied to determine the metabolic activity (MTT) while necrotic effects were assessed by LDH-release measurements. RESULTS: Cactus pear polysaccharides differed predominantly in their glucose and uronic acid content. The filtration of pressed juices altered the amounts of high molecular weight compounds. The proliferation of NHDF and HaCaTs was significantly stimulated by cactus pear polysaccharides and ice plant pressed juices not until 72 h of incubation. McPI significantly increased the proliferation of NHDF and HaCaTs while significant effect of McPII was only observed in case of HaCaT-keratinocytes. A dependence on concentration was not observed. Metabolic activity was neither influenced by McPI nor by McPII independent of incubation time. The HaCaT proliferation was not significantly influenced by low concentrations of cactus pear polysaccharides however it was inhibited by 100 µg/mL NPec. 100 µg/mL of NwPS and 1 µg/mL NPec stimulated the proliferation of fibroblasts. The metabolic activity of NHDF was not affected neither by NPec nor by NwPS. Independent of the used concentration NwPS significantly enhanced the metabolic activity of HaCaTs after 48 h of incubation. CONCLUSIONS: Pressed juices of common ice plant and polysaccharides of cactus pear influenced the cell physiology of human keratinocytes and fibroblasts predominantly in a time-dependent manner. The effect was also be related to the concentration and composition as well as the investigated cell type.

Biochemical, microbiological and phytochemical studies on aqueous-fermented extracts from Atropa belladonna L. Part 1--biochemistry and microbiology.

Extraction methods of fresh plants into aqueous-fermented extracts according to German Homoeopathic Pharmacopoeia (HAB), regulation nos. 33 and 34 were evaluated. In the course of production, the extraction is accompanied by fermentation and the resulting preparation is stored for at least 6 months until further processing. The present work aimed at revealing the underlying biochemical reactions during manufacture and storage. In addition, the responsible microorganisms were isolated and identified. To study the robustness of the preparation method, formulation components as well as production conditions were varied. Additionally, questions were addressed at the reproducibility of the method and a comparison with an ethanolic extract was also performed. From 2006 to 2009, 110 extracts from the fresh flowering herb of Atropa belladonna var. belladonna (L.) were produced and analyzed. The results show that lactic acid bacteria (LAB) are primarily involved in the fermentation process, mainly producing lactic acid besides acetic acid and ethanol. The homofermentative Lactobacillus plantarum and the heterofermentative Lactobacillus brevis were identified as predominant lactic acid bacteria. Finally, factors for a successful fermentation are proposed.

Efficacy of an aqueous Pelargonium sidoides extract against herpesvirus.

The compounds of an aqueous root extract of the African medicinal plant Pelargonium sidoides were analysed by LC-MS spectroscopy and the antiviral effect of this extract against herpes simplex virus was examined in cell culture. Besides predominant coumarins, simple phenolic structures as well as flavonoid and catechin derivatives were identified as major constituents in the Pelargonium extract. The inhibitory activity of this extract against herpes simplex virus type 1 (HSV-1) and herpes simplex virus type 2 (HSV-2) was tested in vitro on RC-37 cells using a plaque reduction assay and exhibited high antiviral activity against both herpesviruses in viral suspension tests. The 50% inhibitory concentration (IC(50)) of the aqueous Pelargonium sidoides extract for herpes simplex virus plaque formation was determined at 0.00006% and 0.000005% for HSV-1 and HSV-2, respectively. At maximum noncytotoxic concentrations of the extract, plaque formation was significantly reduced by more than 99.9% for HSV-1 and HSV-2 and a clear concentration-dependent antiviral activity against HSV could be demonstrated for this extract. In order to determine the mode of antiviral action, the extract was added at different times to the cells or viruses during the infection cycle. Both herpesviruses were significantly inhibited when pretreated with the plant extract or when the extract was added during the adsorption phase, whereas acyclovir demonstrated antiviral activity only intracellularly during replication of HSV. These results indicate that P. sidoides extract affected the virus before penetration into the host cell and reveals a different mode of action when compared to the classical drug acyclovir. Hence this extract is capable of exerting an antiviral effect on herpes simplex virus and might be suitable for topical therapeutic use as antiviral drug both in labial and genital herpes infection.

Comparative in vitro study on the anti-herpetic effect of phytochemically characterized aqueous and ethanolic extracts of Salvia officinalis grown at two different locations.

Aqueous and ethanolic extracts of Salvia officinalis (Lamiaceae) from two different locations (Garden and Swabian Mountains) were examined in vitro on RC-37 cells for their antiviral activity against herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) using a plaque reduction assay. The 50% inhibitory concentrations (IC50) of the extracts for HSV plaque formation were determined in dose-response studies. All extracts tested revealed a high virucidal activity against free HSV-1 and HSV-2. The experimental data exhibited a significant higher sensitivity of HSV against the extracts derived from Garden in comparison with those from Swabian Mountains. The most active one was the Garden 20% ethanol extract with IC50 values of 0.18 microg/ml for HSV-1 and 0.04 microg/ml for HSV-2. In order to identify the mode of antiviral action, the extracts were added to the host cells (RC-37) or viruses at different stages of infection. Independently of the location, both types of herpes viruses were considerably inactivated after treatment with the extracts prior to cell infection. Plaque formation was significantly reduced by >90% for HSV-1 and by >99% for HSV-2. Pretreatment of the host cells with both Garden and Swabian Mountains 20% and 40% ethanolic extracts prior to virus infection revealed a strong reduction of HSV-2 plaque formation by 94% and 70% (Garden) and 99% and 45% (Swabian Mountains), respectively. In time-activity studies with free HSV-1 over a period of 2h, a clearly time-dependent activity was demonstrated whereby the ethanolic extracts of both locations revealed a much higher activity than the aqueous ones. The 20% ethanolic extracts of both locations are of special interest and were effective when added to host cells and free virus. A topical application with a dual mode of action would be ideal against recurrent herpes infections.

Amino acid composition and betaxanthin formation in fruits from Opuntia ficus-indica.

In contrast to earlier reports high levels of taurine (2-aminoethanesulfonic acid) were found in fruit juices of three cultivars of Opuntia ficus-indica (L.) Mill. Whereas the occurrence of taurine in plant tissue was thought to be restricted to algae, fungi, and the endosperm of some higher plants, prickly pear proved to be a rich source of dietary taurine. Using L-taurine as the amino compound, a new betaxanthin was synthesized by partial synthesis. On the basis of chemical and spectral evidence its structure was determined to be the taurine-immonium-conjugate of betalamic acid. Also betalamic acid could be detected in yellow and orange coloured cultivars of Opuntia ficus-indica for the first time. In spite of the high levels of L-taurine accompanied by the occurrence of betalamic acid, the corresponding betaxanthin could not be detected in the fruit tissue.