Apple Pomace as Potential Source of Natural Active Compounds.

Apple pomace is a waste product of the apple manufacturing industry that has been in the focus of life sciences as it represents a low-cost source of fruit-derived compounds. High fruit consumption is associated with beneficial health effects, and therefore, apple pomace and its constituents raise therapeutic interest. The present work reviews (i) the chemical constituents of apple pomace, (ii) optimized extraction methods of apple pomace compounds, and (iii) biological activities of apple pomace. Current evidence of apple pomace influence on digestion and metabolism, cholesterol and triglyceride homeostasis, diabetes, and sex hormones is summarized. Furthermore, studies regarding its antioxidative, anti-inflammatory, antiproliferative, antibacterial and antiviral effects are presented. The review concludes that apple pomace is an underutilized waste product of the apple industry with the potential of being processed for its nutritional and pharmaceutical value.

Sesquiterpenes from Neurolaena lobata and their antiproliferative and anti-inflammatory activities.

Five new sesquiterpenes, neurolobatin A (1), neurolobatin B (2), 5ß-hydroxy-8ß-isovaleroyloxy-9α-hydroxycalyculatolide (3), 3-epi-desacetylisovaleroylheliangine (4), and 3ß-acetoxy-8ß-isovaleroyloxyreynosin (5), were isolated from the aerial parts of Neurolaena lobata. The structures were established by means of a combined spectroscopic data analysis, including ESIMS, APCI-MS, and 1D- and 2D-NMR techniques. Neurolobatin A (1) and B (2) are unusual isomeric seco-germacranolide sesquiterpenes with a bicyclic acetal moiety, compounds 3 and 4 are unsaturated epoxy-germacranolide esters, and compound 5 is the first eudesmanolide isolated from the genus Neurolaena. The isolated compounds (1-5) were shown to have noteworthy antiproliferative activities against human tumor cell lines (A2780, A431, HeLa, and MCF7). The anti-inflammatory effects of 1-5, evaluated in vitro using LPS- and TNF-α-induced IL-8 expression inhibitory assays, revealed that all these compounds strongly down-regulated the LPS-induced production of IL-8 protein, with neurolobatin B (2) and 3-epi-desacetylisovaleroylheliangine (4) being the most effective.

Cytotoxic constituents from Lobaria scrobiculata and a comparison of two bioassays for their evaluation.

Lichens are resilient organisms, known for their unique profiles of secondary metabolites and for exhibiting antioxidative, antibacterial, and cytotoxic effects. Analyzing the cytotoxic potential of Lobaria scrobiculata, a bioassay-guided fractionation strategy yielded seven known metabolites, with two of these compounds, 2 and 3, exhibiting cytotoxicity against HL-60 cells. In order to verify the potential impact of degradation on observed bioactivity, a purity and stability evaluation was conducted. The consistency of results obtained by the water-soluble tetrazolium salt-1 assay and trypan blue cytotoxicity assay was evaluated for selected compounds.

Influence of vinegar and wine processing on the alkaloid content and composition of the traditional Chinese medicine Corydalis Rhizoma (Yanhusuo).

Corydalis Rhizoma is the dried tuber of Corydalis yanhusuo W.T. Wang which is used in traditional Chinese medicine for pain relief and blood activation. Before being used in the clinics, C. yanhusuo is traditionally processed through dry-frying or frying with vinegar, wine or salt. In this study, eleven alkaloids from Corydalis Rhizoma, namely protopine (1), α-allocryptopine (2), tetrahydrocolumbamine (3), coptisine (4), palmatine (5), berberine (6), dehydrocorydaline (7), D,L-tetrahydropalmatine (8), tetrahydroberberine (9), corydaline (10) and tetrahydrocoptisine (11) were simultaneously quantified using a newly developed high performance liquid chromatography-diode array detector (HPLC-DAD) method. The influence of vinegar and wine processing on the content of the main alkaloids of Corydalis Rhizoma was investigated. For this purpose, two common formulations with clinical application, namely the water decoction of Corydalis Rhizoma and its formula Jin Ling Zi San (combination of Corydalis Rhizoma and Toosendan Fructus) were studied. In the two water decoctions, wine and vinegar processing increased the amount of tertiary alkaloids. The differences were more pronounced for Jin Ling Zi San, in which case the content of all tertiary alkaloids (compounds 1, 2, 3, 8, 9, 10, 11) was increased by wine processing.

Hellebrin and its aglycone form hellebrigenin display similar in vitro growth inhibitory effects in cancer cells and binding profiles to the alpha subunits of the Na+/K+-ATPase.

BACKGROUND: Surface-expressed Na+/K+-ATPase (NaK) has been suggested to function as a non-canonical cardiotonic steroid-binding receptor that activates multiple signaling cascades, especially in cancer cells. By contrast, the current study establishes a clear correlation between the IC50in vitro growth inhibitory concentration in human cancer cells and the Ki for the inhibition of activity of purified human α1ß1 NaK. METHODS: The in vitro growth inhibitory effects of seven cardiac glycosides including five cardenolides (ouabain, digoxin, digitoxin, gitoxin, uzarigenin-rhamnoside, and their respective aglycone forms) and two bufadienolides (gamabufotalin-rhamnoside and hellebrin, and their respective aglycone forms) were determined by means of the MTT colorimetric assay and hellebrigenin-induced cytotoxic effects were visualized by means of quantitative videomicroscopy. The binding affinity of ten of the 14 compounds under study was determined with respect to human α1ß1, α2ß1 and α3ß1 NaK complexes. Lactate releases and oxygen consumption rates were also determined in cancer cells treated with these various cardiac glycosides. RESULTS: Although cardiotonic steroid aglycones usually display weaker binding affinity and in vitro anticancer activity than the corresponding glycoside, the current study demonstrates that the hellebrin / hellebrigenin pair is at odds with respect to this rule. In addition, while some cardiac steroid glycosides (e.g., digoxin), but not the aglycones, display a higher binding affinity for the α2ß1 and α3ß1 than for the α1ß1 complex, both hellebrin and its aglycone hellebrigenin display ~2-fold higher binding affinity for α1ß1 than for the α2ß1 and α3ß1 complexes. Finally, the current study highlights a common feature for all cardiotonic steroids analyzed here, namely a dramatic reduction in the oxygen consumption rate in cardenolide- and bufadienolide-treated cells, reflecting a direct impact on mitochondrial oxidative phosphorylation. CONCLUSIONS: Altogether, these data show that the binding affinity of the bufadienolides and cardenolides under study is usually higher for the α2ß1 and α3ß1 than for the α1ß1 NaK complex, excepted for hellebrin and its aglycone form, hellebrigenin, with hellebrigenin being as potent as hellebrin in inhibiting in vitro cancer cell growth.

Structure-activity relationship analysis of bufadienolide-induced in vitro growth inhibitory effects on mouse and human cancer cells.

The in vitro growth inhibitory effects of 27 bufadienolides and eight degradation products, with two cardenolides (ouabain and digoxin) chosen as reference compounds, were analyzed by means of an MTT colorimetric assay in six human and two mouse cancer cell lines. A structure-activity analysis was then performed to highlight the most important substituents relating to the in vitro growth inhibitory activity of bufadienolides in cancer cells. Thus, the current study revealed that various bufadienolides, including gamabufotalin rhamnoside (1a), bufotalin (2a), and hellebrin (3a), displayed higher growth inhibitory activities for various human cancer cell lines when compared to ouabain and digoxin. Gamabufotalin rhamnoside (1a) was the only compound that displayed growth inhibitory effects of <1 µM in mouse cancer cells that expressed mutated forms of the Na(+),K(+)-ATPase α-1 subunit. In addition, all genins and degradation products displayed weaker (if any) in vitro growth inhibitory effects on cancer cells when compared to their respective glycosylated homologue, with the exception of hellebrigenin (3b), which was as active as hellebrin (3a).

GABA(A) receptor modulators from the Chinese herbal drug Junci Medulla--the pith of Juncus effusus.

The gamma-amino butyric acid (GABA) type A (GABA(A)) receptor represents a crucial target for clinical agents in the treatment of anxiety and insomnia. Using the two-microelectrode voltage clamp technique on recombinant α1ß2γ(2S) GABA (A) receptors, effusol (1) and dehydroeffusol (2) were isolated in a bioactivity-guided approach from the pith of Juncus effusus L. Both compounds concentration-dependently enhanced GABA induced chloride currents (I(GABA)) by a maximum 188 ± 20 (1) and 239 ± 18 % (2), independent of the benzodiazepine (BZ) binding site. This activity on the GABA (A) receptor may explain the traditional use of J. effusus as a sedative and anxiolytic agent in Chinese medicine.

Towards modernization of the formulation of the traditional uighur medicine herbal preparation abnormal savda munziq.

Abnormal Savda Munziq (ASMq) is a herbal preparation used in Traditional Uighur Medicine for the treatment and prevention of diabetes, cardiovascular diseases, chronic asthma and cancer. The recommended dose of this decoction for cancer patients is 500 mL administered orally three times a day. Our approach aimed at reducing the high amount of fluid intake required by fractionation of ASMq guided by the antiproliferative activity on HL-60 cells. The fractionation of ASMq resulted in the preparation of an active extract, Extr-4. Using solid phase extraction, Extr-4 was further fractionated into five fractions (SPE-0, SPE-20, SPE-40, SPE-60 and SPE-80), with SPE-40 showing the strongest antiproliferative activity. Caffeic acid, rutin, isoquercitrin, isorhamnetin 3-O-rutinoside, apigenin 7-O-glucoside, rosmarinic acid, luteolin and formononetin were identified in Extr-4 and fractions thereof by means of TLC, HPLC-DAD and LC-MS. SPE-40 contained the main compounds responsible for the antiproliferative activity on HL-60 cells. Thus, a phenolic fraction with high antiproliferative activity on HL-60 cells was obtained from ASMq through the bioassay-guided fractionation process. This could provide a better pharmaceutical formulation that minimizes the administration inconveniencies of a high volume (1.5 L per day) of ASMq decoction for cancer patients.

Fractionation of an Extract of Pluchea odorata Separates a Property Indicative for the Induction of Cell Plasticity from One That Inhibits a Neoplastic Phenotype.

Introduction. Several studies demonstrated that anti-inflammatory remedies exhibit excellent anti-neoplastic properties. An extract of Pluchea odorata (Asteraceae), which is used for wound healing and against inflammatory conditions, was fractionated and properties correlating to anti-neoplastic and wound healing effects were separated. Methods. Up to six fractionation steps using silica gel, Sephadex columns, and distinct solvent systems were used, and eluted fractions were analysed by thin layer chromatography, apoptosis, and proliferation assays. The expression of oncogenes and proteins regulating cell migration was investigated by immunoblotting after treating HL60 cells with the most active fractions. Results. Sequential fractionations enriched anti-neoplastic activities which suppressed oncogene expression of JunB, c-Jun, c-Myc, and Stat3. Furthermore, a fraction (F4.6.3) inducing or keeping up expression of the mobility markers MYPT, ROCK1, and paxillin could be separated from another fraction (F4.3.7), which inhibited these markers. Conclusions. Wound healing builds up scar or specific tissue, and hence, compounds enhancing cell migration support this process. In contrast, successful anti-neoplastic therapy combats tumour progression, and thus, suppression of cell migration is mandatory.

Comparative evaluation of two different Artemisia dracunculus L. cultivars for blood sugar lowering effects in rats.

Recent concerns about the potential carcinogenicity of estragole and methyleugenol led a number of regulatory bodies to call for restrictions on the use of herbs that contain these constituents. A number of medicinal plants produce essential oils that contain estragole and methyleugenol, including Artemisia dracunculus L. (tarragon). Previous studies have proven the antidiabetic properties of tarragon. In order to address the safety concerns of estragole containing tarragon extracts, an extraction procedure was developed to minimize the estragole and methyleugenol content in tarragon extracts and the ethanol versus aqueous extracts from two Artemisia dracunculus cultivars (French and Russian tarragon) were tested for blood glucose lowering effects in rats. It could be demonstrated that aqueous extracts of both Artemisia cultivars did not contain detectable amounts of estragole and methyleugenol, whereas ethanol extracts (60% v/v) of the French cultivar contained higher levels of the aforementioned compounds than those of the Russian cultivar. Further testing revealed that Russian tarragon lowered blood glucose levels in rats after glucose challenge, with the ethanol extract being as active as the aqueous extract. The results suggest that by using adequate production procedures the amount of potentially harmful compounds in extracts can be limited without affecting the overall pharmacological activities of these preparations.

Ferruginenes A-C from Rhododendron ferrugineum and their cytotoxic evaluation.

Three new compounds, ferruginenes A (1) and B (2) and a mixture of C-5'(R) and C-5'(S) ferruginene C (3) diastereomers, have been isolated from a cytotoxic chloroform-soluble fraction of the leaves of Rhododendron ferrugineum together with 12 known compounds. The structures of these new compounds were elucidated by analyses of NMR spectroscopic and mass spectrometric data. Compounds 1-3 were tested for their cytotoxicity against three human cancer cell lines, namely, HL-60, HeLa-S3, and MCF-7.

Identification of ostruthin from Peucedanum ostruthium rhizomes as an inhibitor of vascular smooth muscle cell proliferation.

Inhibition of vascular smooth muscle cell (VSMC) proliferation is of substantial interest in combating cardiovascular disease. A dichloromethane extract from the rhizomes of Peucedanum ostruthium, a traditionally used Austrian medicinal plant with anti-inflammatory properties, was examined for a putative antiproliferative activity in rat aortic VSMC. This extract inhibited serum (10%)-induced VSMC proliferation concentration dependently. Further identification and biological testing of its major constituents revealed that the coumarin ostruthin (7) is the major antiproliferative substance. In summary, a new bioactivity of P. ostruthium rhizomes is described, and 7 has been identified as the responsible compound.

Ikarugamycin induces DNA damage, intracellular calcium increase, p38 MAP kinase activation and apoptosis in HL-60 human promyelocytic leukemia cells.

Ikarugamycin (IKA) is an antibiotic with strong antiprotozoal and cytotoxic activity. The purpose of our work was to provide insight into the mechanism of action characterizing the cytotoxic effect of IKA in HL-60 leukemia cells in order to evaluate its potential as an antineoplastic agent. Cell viability was reduced in response to IKA (IC(50) of 221.3nM), while the amount of HL-60 cells with a subdiploid DNA content increased significantly after 24h. Apoptotic cell death was confirmed by the cleavage of caspase-9, -8 and -3 using immunoblotting. Single cell gel electrophoresis pointed to an early genotoxic effect. Monitoring of intracellular calcium ([Ca(2+)](i)) levels by flow cytometric analysis of Fluo-3-AM fluorescence indicated an increase in cytosolic calcium that correlated with the cleavage of caspases. In addition, IKA triggered the activation of p38 MAP kinase which was partly dependent on elevated [Ca(2+)](i) concentrations and contributed to caspase activation. The data demonstrate that IKA induced apoptosis in HL-60 cells through genotoxicity and caspase activation which was in part correlated to an increase in intracellular calcium levels and activation of p38 MAP kinase.

Young people's trauma-related cognitions before and after cognitive processing therapy for post-traumatic stress disorder.

OBJECTIVES: Cognitive processing therapy (CPT) is a psychotherapy for post-traumatic stress disorder (PTSD) with a broad evidence base. Change in trauma-related cognitions is considered its primary working mechanism. When trying to integrate a traumatic event into existing cognitive schemas, the adaptive mechanism is changing the schema (accommodation). However, PTSD patients frequently either change their schemas too much (over-accommodation), or cognitively distort the event (assimilation). We aimed to test the hypothesized connections between the three types of cognition and symptom load. DESIGN: This study adds to the literature using 'impact statements', essays on their trauma-related thoughts written by patients at the beginning and end of CPT, to investigate cognitive change and its relationship to symptomatic outcome. METHODS: We analysed statements written by 31 adolescents and young adults who received developmentally adapted CPT (a longer treatment where CPT is the core component) in a randomized controlled trial. RESULTS: As expected, post-CPT statements contained more accommodated and fewer over-accommodated and assimilated clauses than pre-CPT statements. Correlations between cognition frequencies and concurrent symptom load were as expected for assimilation, and, in part, over-accommodation and accommodation. Decreased PTSD and depressive symptoms were correlated with increased accommodated thoughts. For over-accommodation and assimilation, however, expected correlations could not be shown. CONCLUSIONS: Our results support the notion that cognitive change is an important mechanism of change in CPT in a sample of younger, non-English-speaking clients.

Rapid structural identification of cytotoxic bufadienolide sulfates in toad venom from Bufo melanosticus by LC-DAD-MS(n) and LC-SPE-NMR.

Toad venom, namely, "Chansu" in China, has been widely used for the treatment of heart failure, sores, pains, and various cancers. Upon LC-MS analysis of the venom from Bufo melanosticus collected in Indonesia, new bufadienolide sulfates were identified. For a complete characterization, the MeOH extract of the toad venom from B. melanosticus was fractionated by preparative HPLC, and the structures of five new buadienolide sulfates (1-5) along with one new bufogenin (6) were rapidly elucidated on the basis of LC-DAD-MS(n) and LC-SPE-NMR data. The in vitro growth inhibitory activity of these six compounds along with hellebrin (positive control) has been assayed by means of the MTT colorimetric assay in four human and two mouse cancer cell lines. Compound 3 and hellebrin displayed similar and marked in vitro cytotoxicity.

Berberine and a Berberis lycium extract inactivate Cdc25A and induce alpha-tubulin acetylation that correlate with HL-60 cell cycle inhibition and apoptosis.

Berberis lycium Royle (Berberidacea) from Pakistan and its alkaloids berberine and palmatine have been reported to possess beneficial pharmacological properties. In the present study, the anti-neoplastic activities of different B. lycium root extracts and the major constituting alkaloids, berberine and palmatine were investigated in p53-deficient HL-60 cells. The strongest growth inhibitory and pro-apoptotic effects were found in the n-butanol (BuOH) extract followed by the ethyl acetate (EtOAc)-, and the water (H(2)O) extract. The chemical composition of the BuOH extract was analyzed by TLC and quantified by HPLC. 11.1 microg BuOH extract (that was gained from 1mg dried root) contained 2.0 microg berberine and 0.3 microg/ml palmatine. 1.2 microg/ml berberine inhibited cell proliferation significantly, while 0.5 microg/ml palmatine had no effect. Berberine and the BuOH extract caused accumulation of HL-60 cells in S-phase. This was preceded by a strong activation of Chk2, phosphorylation and degradation of Cdc25A, and the subsequent inactivation of Cdc2 (CDK1). Furthermore, berberine and the extract inhibited the expression of the proto-oncogene cyclin D1. Berberine and the BuOH extract induced the acetylation of alpha-tubulin and this correlated with the induction of apoptosis. The data demonstrate that berberine is a potent anti-neoplastic compound that acts via anti-proliferative and pro-apoptotic mechanisms independent of genotoxicity.

Root colonization by symbiotic arbuscular mycorrhizal fungi increases sesquiterpenic acid concentrations in Valeriana officinalis L.

In some medicinal plants a specific plant-fungus association, known as arbuscular mycorrhizal (AM) symbiosis, increases the levels of secondary plant metabolites and/or plant growth. In this study, the effects of three different AM treatments on biomass and sesquiterpenic acid concentrations in two IN VITRO propagated genotypes of valerian ( VALERIANA OFFICINALIS L., Valerianaceae) were investigated. Valerenic, acetoxyvalerenic and hydroxyvalerenic acid levels were analyzed in the rhizome and in two root fractions. Two of the AM treatments significantly increased the levels of sesquiterpenic acids in the underground parts of valerian. These treatments, however, influenced the biomass of rhizomes and roots negatively. Therefore this observed increase was not accompanied by an increase in yield of sesquiterpenic acids per plant. Furthermore, one of the two genotypes had remarkably high hydroxyvalerenic acid contents and can be regarded as a hydroxyvalerenic acid chemotype.