Cellular Stress and p53-Associated Apoptosis by Juniperus communis L. Berry Extract Treatment in the Human SH-SY5Y Neuroblastoma Cells.

Plant phenolics have shown to activate apoptotic cell death in different tumourigenic cell lines. In this study, we evaluated the effects of juniper berry extract (Juniperus communis L.) on p53 protein, gene expression and DNA fragmentation in human neuroblastoma SH-SY5Y cells. In addition, we analyzed the phenolic composition of the extract. We found that juniper berry extract activated cellular relocalization of p53 and DNA fragmentation-dependent cell death. Differentially expressed genes between treated and non-treated cells were evaluated with the cDNA-RDA (representational difference analysis) method at the early time point of apoptotic process when p53 started to be activated and no caspase activity was detected. Twenty one overexpressed genes related to cellular stress, protein synthesis, cell survival and death were detected. Interestingly, they included endoplasmic reticulum (ER) stress inducer and sensor HSPA5 and other ER stress-related genes CALM2 and YKT6 indicating that ER stress response was involved in juniper berry extract mediated cell death. In composition analysis, we identified and quantified low concentrations of fifteen phenolic compounds. The main groups of them were flavones, flavonols, phenolic acids, flavanol and biflavonoid including glycosides of quercetin, apigenin, isoscutellarein and hypolaetin. It is suggested that juniper berry extract induced the p53-associated apoptosis through the potentiation and synergism by several phenolic compounds.

Characterization of volatile and semi-volatile compounds in green and fermented leaves of Bergenia crassifolia L. by gas chromatography-mass spectrometry and ID-CUBE direct analysis in real time-high resolution mass spectrometry.

Chemical compositions of volatile and semi-volatile components in green and fermented leaves of Bergenia crassifolia L. were studied. Leaf components were identified using gas chromatography with low resolution mass spectrometry and direct analysis in real time (DART) high resolution mass spectrometry with an ID-CUBE ion source. Phytol, nerolidol, geraniol, linalool, alpha-bisabolol, alpha-bisabololoxide B, alpha-cadinol, delta-cadinene, alpha-terpineol and several other marker compounds of special interest were defined, for which the process of fermentation significantly changed their content in the leaves. Low resolution El GC-MS and ID-CUBE DART-HRMS were found to be complementary methods, as they provide different information, helpful to increase the confidence of identification.

Rapid profiling of phenolic compounds of green and fermented Bergenia crassifolia L. leaves by UPLC-DAD-QqQ-MS and HPLC-DAD-ESI-QTOF-MS.

Bergenia crassifolia L., Saxifragaceae, is an evergreen perennial plant known in traditional medicine of Russia, Mongolia and China. Polyphenols are responsible for the number of pharmacological effects of Bergenia. UPLC-DAD-QqQ-MS and LC-DAD-ESI-QTOF-MS were used for the rapid profiling of phenolic compounds, mainly hydrolysable tannins. Green leaves consisted of 55% ellagitannins, 29% gallic acid derivatives and 11% flavonoids, with the remaining gallic acid, arbutin, bergenin and caffeoyl quinic acid. In fermented leaves, 31% of gallic acid was found, followed with 28% ellagitannins, 18% gallic acid derivatives and 18% flavonoids, with the remaining caffeoyl quinic acid, bergenin and arbutin. Tellimagrandin I, pedunculagin, caffeoyl quinic acid, monogalloyl quinic acid, 1-O-galloylglucose and 1,2,6-tri-O-galloylglucose were identified for the very first time.

A water-alcohol extract of Citrus grandis whole fruits has beneficial metabolic effects in the obese Zucker rats fed with high fat/high cholesterol diet.

Epidemiological studies suggest that citrus fruits and compounds such as flavonoids, limonoids and pectins have health promoting effects. Our aim was to study the effects of Citrus grandis (L.) Osbeck var. tomentosa hort. fruit extract on the energy metabolism. A whole fruit powder from dry water and alcohol extracts of C. grandis containing 19% naringin flavonoid was prepared. The effects of the citrus extract were followed in the obese Zucker rats fed with the HFD. The circulatory levels of GLP-1 decreased significantly by the extract in comparison to the HFD group, whereas the decreased ghrelin levels were reversed. The levels of PYY were decreased in all HFD groups. The leptin amounts decreased but not significantly whereas insulin and amylin were unchanged. The cholesterol and glucose levels were somewhat but not systematically improved in the HFD fed rats. Further studies are needed to identify the active compounds and their mechanisms.

Effect of Bergenia crassifolia L. extracts on weight gain and feeding behavior of rats with high-caloric diet-induced obesity.

The objective of this study was to evaluate the feeding behavior and weight gain in rats with high-calorie diet-induced obesity that are treated with Bergenia crassifolia black and fermented leaves extracts. The daily dietary intake of all treated animals was reduced to 40% compared with the control group on day 22 of the experiment. A significant improvement in glucose tolerance was noted after 7 days of treatment with the Bergenia extracts. In rats treated with an extract of black leaves for 7 days, a significant reduction in the serum triglyceride level, 45% (p<0.05), compared with the control group was observed. However, the treatment did not affect the cholesterol level. Our results provide evidence for the potential use of B. crassifolia as an appetite and energy intake suppressant.

Antioxidant, pro-oxidant and cytotoxic properties of parsley.

Parsley (Petroselinum crispum) leaves were macerated with a mixture of methanol: water: acetic acid to produce a crude extract which was then defatted with (40°-60°) petrol. Antioxidant activity of the extract was evaluated using a battery of in vitro assays, viz., iron(iii) reduction, iron(ii) chelation and free radical scavenging assays. Evaluation of the pro-oxidant activity of the extract was based upon its effects upon DNA fragmentation and protein carbonylation. Cytotoxicity and apoptotic effects of the extract were determined in non-cancerous CV1-P fibroblast and cancerous A375 melanoma cells using MTT and LDH tests and caspase 3-like activity assay. The highest concentration, 2.0 mg ml(-1), decreased the viability of both cell lines, however, the cancerous melanoma cells were slightly susceptible to the effects. The observed cytotoxicity was not due to the caspase 3 activity. In conclusion, the toxicity might be explained by the pro-oxidative activity of components within the extract against proteins and/or DNA but it is not related to caspase 3-dependent apoptosis within cells.

Antioxidant and pro-oxidant evaluation of a Potentilla alba L. rhizome extract.

Using spectrophotometric methods, a H(2) O-soluble Potentilla alba L. rhizome extract was evaluated phytochemically, i.e., the total phenol, flavonoid, flavonol, flavanone, and proanthocyanidin contents were determined, and its antioxidant and pro-oxidant properties, i.e., the Fe(III) reductive and the Fe(II) chelating properties, the 1,1-diphenyl-2-picrylhydrazyl radical (DPPH(*)), N,N-dimethyl-p-phenylenediamine (DMPD(*+)), and superoxide anion radical (O2*-)-scavenging activities, the capacity to inhibit hydroxyl radical (HO(*))-mediated deoxy-D-ribose and phospholipid degradation, and the interaction with the Cu-catalyzed HO(*) -mediated DNA degradation, were determined. The extract was found to contain a range of phenolic compounds recognized to possess strong antioxidant-like properties. Moreover, the extract demonstrated dose-dependent activities in all the antioxidant assays with the exception of the DNA-degradation assay, where the components within the extract interfered with the assay components at concentrations ≥1.00 mg/ml. Potentilla species are known for their curative properties, with aerial/subterranean parts being prescribed for numerous indications. The data presented here suggests, though does not conclude, that the rhizomes contain compounds possessing a range of antioxidant-related properties, which may underpin the therapeutic, viz., anti-inflammatory and adaptogenic effects, ascribed to species of this genus.

Iron(III) reducing and antiradical activities of three Sideritis from Turkey.

CONTEXT: Sideritis species (Lamiaceae) are widely used as herbal tea and have been used in folk medicine for their anti-inflammatory, anti-rheumatic, digestive, and antimicrobial activities in Turkey. Sideritis dichotoma Huter., Sideritis erythrantha Boiss. var. cedrotorum, and Sideritis vuralii H. Duman et Baser are available as commercial products in Turkey. OBJECTIVE: The antiradical activities of the various solvent extracts of Sideritis species are investigated here for the first time. MATERIALS AND METHODS: Plant samples were sequentially extracted with n-hexane, dichloromethane, methanol, and aqueous methanol (50%, v/v) in Soxhlet apparatus. The extracts of Camellia sinensis (L.) Kuntze (Theaceae) were also prepared for use as a positive control. Total phenolics, iron(III) reductive effects, and 1,1-diphenyl-2-picrylhydrazyl (DPPH•) radical scavenging activities of the all extracts were measured colorimetrically. RESULTS: The aqueous MeOH and MeOH extracts contained the highest amount of total phenols, whereas the n-hexane extract contained the lowest amounts. The polar extracts of C. sinensis showed higher antiradical activity and also iron(III) reductive effects than the Sideritis species; however, the non-polar extracts of Sideritis species were found to be more active than those from C. sinensis in the iron(III) reductive assay and in the DPPH(•) assay as well. But none of the extracts was found to be as active as with positive controls, viz., ascorbic acid, butylated hydroxyanisole (BHA), and Trolox. DISCUSSION AND CONCLUSION: These results can be shown to have antioxidant activities of these Sideritis species and support the ethnopharmacological use of these Sideritis plants.

Salvia officinalis L.: composition and antioxidant-related activities of a crude extract and selected sub-fractions.

The composition and antioxidant properties of a methanol: acetic acid (99:1, v/v) soluble crude extract isolated from S. officinalis L. leaves through maceration and selected fractions isolated thereof are presented in this study. The total phenol content was estimated as gallic acid equivalents, whilst qualitative-quantitative phenolic content was determined using high performance liquid chromatography with photodiode array detection. Antioxidant evaluation consisted of ferric reductive capacity and 1,1-diphenyl-2-picrylhydrazyl and hydroxyl free radical scavenging determinations. The crude extract contained hydroxybenzoic acids, hydroxycinnamic acids, flavonoids and diterpenoids, whilst caffeic acid, carnosic acid, luteolin, luteolin-7-O-glucoside and rosmarinic acid were identified from their chromatographic and spectral characteristics and quantified from their respective calibration curves. The crude extract and sub-fractions demonstrated varying degrees of efficacy in the antioxidant-related assays used, except the n-hexane fraction, which was unable to reduce iron(III) at reasonable concentrations. Although the positive controls, ascorbic acid, BHA and BHT, were more potent than the S. officinalis samples, two fractions were significantly (p < 0.05) more potent iron(III) reducing agents than pycnogenol, a proanthocyanidin-rich commercial preparation.

Ocimum basilicum L.: phenolic profile and antioxidant-related activity.

Ocimum basilicum L. leaf material was extracted by maceration with (80:20:1 v/v/v) methanol: water: acetic acid to produce a crude extract (CE), which was further fractionated by liquid-liquid extraction to isolate light petroleum (PE), ethyl acetate (EtOAc), n-butanol (n-BuOH) and H2O-soluble sub-fractions. The total phenol and flavonoid contents of the resulting samples were estimated using colorimetric-based methods, and their iron(III) reductive and free radical scavenging activities were determined in a battery of in vitro assays. The CE and sub-fractions contained phenolic compounds and flavonoids. The samples, except for PE, gave a positive result for the presence of flavones and flavonols; however, flavanones only appeared to be present in the CE. In iron(III) reduction, CE and n-BuOH were the most potent followed by EtOAc and H2O (statistically indistinguishable, p > 0.05). However, in the ferric reducing antioxidant power assay, H2O was the most potent followed by CE and EtOAc (statistically indistinguishable, p > 0.05) and n-BuOH and PE. In 1,1-diphenyl-2-picrylhydrazyl scavenging, all the samples, except PE, were effective against this reactive nitrogen species, with CE, EtOAc and n-BuOH being the most potent (statistically indistinguishable, p > 0.05). In alkylperoxyl scavenging, all the samples, except for PE, were effective against this reactive oxygen species (ROS). In superoxide anion scavenging, all the samples were capable of scavenging this ROS with CE being the most effective, followed by n-BuOH and H2O (statistically indistinguishable, p > 0.05) and EtOAc and PE. Similarly, in hydroxyl scavenging, all the samples were capable of scavenging this ROS with CE and n-BuOH being the most effective (statistically indistinguishable, p > 0.05) followed by EtOAc and H2O (statistically indistinguishable, p > 0.05) and PE.

Nanodispersions of taxifolin: impact of solid-state properties on dissolution behavior.

Nanosizing is an advanced formulation approach to address the issues of poor aqueous solubility of active pharmaceutical ingredients. Here we present a procedure to prepare a nanoparticulate formulation with the objective to enhance dissolution kinetics of taxifolin dihydrate, a naturally occurring flavonoid with antioxidant, anti-inflammatory, and hepatoprotective activities. Polyvinylpirrolidone was selected as a carrier and the solid nanodispersions of varying compositions were prepared by a co-precipitation technique followed by lyophilization. The formulation technology reported herein resulted in aggregate-free, spherical particles with the mean size of about 150 nm, as observed by scanning electron microscopy and measured by photon correlation spectroscopy. Furthermore, the co-precipitation process caused taxifolin dihydrate to convert into an amorphous form as verified by X-ray powder diffraction, differential scanning calorimetry, hot stage microscopy and Raman spectroscopy. Finally, in vitro dissolution behavior of the nanodispersion of taxifolin was shown to be superior to that of either pure drug or a drug-polymer physical mixture, reaching 90% of taxifolin released after 30 min. Such enhanced drug release kinetics from the nanodispersion was attributed to both the reduced particle size and the loss of crystallinity.

Phenolic profile and antioxidant evaluation of Mentha x piperita L. (peppermint) extracts.

Seven extracts were prepared from Mentha x piperita (peppermint) leaves in sequence using a Soxhlet apparatus, viz. (40-60 degrees) light petroleum (PE), dichloromethane (CH2Cl2), acetonitrile (ACN), ethyl acetate (EtOAc), methanol (MeOH), n-butanol and water (H2O) extracts. The phenolic and flavonoid content of each extract were estimated using spectrophotometric methods whilst a qualitative-quantitative analysis was made by reverse-phase high performance liquid chromatography coupled with photodiode array detection (HPLC-PDA). Each extract was assessed in a battery of six antioxidant-related assays so as to determine their iron(III) reductive, iron(II) chelating and free radical scavenging abilities. The MeOH-soluble extract contained the greatest content of total phenols and flavonoids based upon the Folin-Ciocalteu and 2-aminoethyl diphenylborate reagent data and HPLC-PDA analysis. Based upon the chromatographic and UV-spectral data, the leaves principally contained the cinnamic acid caffeic acid, the depside rosmarinic acid and flavonoids (flavones and flavanones). Eriocitrin (383.3 +/- 2.2 mg/g extract) and rosmarinic acid (381.2 +/- 1.9 mg/g extract) were the most abundant components identified within the leaves, whilst naringenin-7-O-glucoside (0.8 +/- 0.01 mg/g extract) was the least abundant component identified being found only in the EtOAc-soluble extract. The EtOAc, ACN and H2O-soluble extracts demonstrated the most potent iron(III) reductive and 1,1'-diphenyl-2-picrylhydrayl, 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonate) and hydroxyl free radical scavenging properties; however, the H2O and CH2Cl2-soluble extracts were the most potent extracts in the beta-carotene-linoleic acid bleaching inhibition assay. In terms of iron(II) chelation--an important antioxidant property--the PE, MeOH and H2O extracts demonstrated moderate iron(II) chelating activity.

Metabolism of berry anthocyanins to phenolic acids in humans.

We studied the metabolism of berry anthocyanins to phenolic acids in six human subjects by giving them bilberry-lingonberry puree with and without oat cereals. Puree + cereals contained 1435 micromol of anthocyanins and 339 micromol of phenolic acids. The urinary excretion of measured 18 phenolic acids increased 241 micromol during the 48 h follow-up after the puree + cereals supplementation. The excretion peak of dietary phenolic acids was observed at 4-6 h after the puree + cereals supplementation and 2 h earlier after the supplementation of the puree alone. Homovanillic and vanillic acids were the most abundant metabolites, and they were partly produced from anthocyanins. No gallic acid, a fragmentation product of delphinidin glycosides, was detected, and only a very low amount of malvidin glycosides was possibly metabolized to syringic acid. Although anthocyanins were partly fragmented to phenolic acids, still a large part of metabolites remained unknown.

Chemical and antioxidant evaluation of Indian gooseberry (Emblica officinalis Gaertn., syn. Phyllanthus emblica L.) supplements.

Indian gooseberry (Emblica officinalis Gaertn.) (Euphorbiaceae) has a distinguished history in Ayurveda medicine and is ascribed a number of medicinal properties and as a dietary supplement, its use is increasing in Western countries. It is thought that its beneficial properties are a function of its antioxidant potency. The study investigated the chemistry and antioxidant properties of four commercial E. officinalis fruit extracts in order to determine if there are any qualitative-quantitative differences. All extracts produced positive responses in the total phenol, total flavonoid and total tannin assays. The presence of predominantly (poly)phenolic analytes, e.g. ellagic and gallic acids and corilagin, was confirmed by RP-HPLC coupled with photodiode array detection. Despite ascorbic acid being a major constituent of E. officinalis fruits, the furanolactone could not be identified in one of the samples. The extracts demonstrated varying degrees of antioxidative efficacy. The extract designated IG-3 was consistently amongst the most effective extracts in the iron(III) reduction and 1,1-diphenyl-2-picrylhydrazyl and superoxide anion radical scavenging assays while the extract designated IG-1 demonstrated the best hydroxyl radical scavenging activity. All extracts appeared to be incapable of chelating iron(II) at realistic concentrations.

Acetylcholinesterase inhibitory guided fractionation of Melissa officinalis L.

The plant Melissa officinalis L. has been used traditionally in the treatment of cognitive dysfunction. Based on its traditional medicinal use, it was assessed for its clinical efficacy in mild to moderate Alzheimer's patients. The plant was effective in the management of the disease. Therefore, based on this result, a similar plant extract was prepared in order to be screened for bioactivities which are relevant in Alzheimer's disease therapy. The extract was recently screened for antioxidant activity and it showed a wide range of antioxidant properties. Another important bioactivity is acetylcholinesterase inhibition, which the extract was screened for in the current investigation. The extract was capable of inhibiting the enzyme in a time and dose-dependent manner. Activity of the extract at 10 min was estimated as 1.72+/-0.16 microg equivalents of physostigmine/mg of the extract. Acetylcholinesterase inhibitory guided fractionation of the extract was then carried out. Most of the fractions showed inhibitory activity and were more potent than the extract. The contents of the most potent fraction were identified as cis- and trans-rosmarinic acid isomers and a rosmarinic acid derivative using LC-DAD-ESI-MS and NMR methods.

Preventing airborne infection with an intranasal cellulose powder formulation (Nasaleze travel).

A total of 52 volunteers were recruited to take part in a dual-centered, randomized, blinded study so investigators could determine whether the level of airborne infection could be significantly reduced in patients randomly assigned to treatment with either Nasaleze cellulose extract alone or a combination of Nasaleze cellulose and powdered garlic extract (PGE). One puff into each nostril was recommended, and volunteers who developed an infection while traveling were told to use at least 3 puffs per nostril until symptoms were reduced. This study took place over an 8-wk period across Finland and the United Kingdom between November 2006 and March 2007. Volunteers were instructed to use a 5-point scale to assess their health and to record infectious episodes and symptoms in a daily diary. The activetreatment group (Nasaleze cellulose with PGE) experienced significantly fewer infections than the control group (20 vs 57; P<.001) and far fewer days on which an infection was obviously present (126 d in the active group vs 240 d in the control group; P<.05). Consequently, volunteers in the active group were less likely to pick up an airborne infection when PGE was added to this novel cellulose extract. Volunteers in the control group were much more likely to report more than 1 infectious episode over the treatment period or to endure longer periods of infection. The investigators concluded that the combination Nasaleze Travel formulation significantly reduced the number of airborne infections to which volunteers were exposed while traveling.

Consumption of juice fortified with oregano extract markedly increases excretion of phenolic acids but lacks short- and long-term effects on lipid peroxidation in healthy nonsmoking men.

Oregano has been shown to possess antioxidant capacity in various in vitro models and has thus been suggested to be potentially beneficial to human health, but studies in humans are lacking. The aim of this study was to investigate the bioavailability and the effects of Origanum vulgare extract supplementation on serum lipids and lipid peroxidation in healthy nonsmoking men. A four-week double-blinded supplementation trial was concluded in which volunteers (n = 45) were randomized to consume daily mango-orange juice (placebo), mango-orange juice enriched with 300 mg/d total phenolic compounds from oregano extract, or mango-orange juice enriched with 600 mg/d total phenolic compounds from oregano extract. The excretion of phenolic compounds was markedly increased in the higher phenolic group as compared to the placebo group, but no significant changes were observed in the safety parameters, serum lipids, or biomarkers of lipid peroxidation.

Ingestion of oregano extract increases excretion of urinary phenolic metabolites in humans.

Despite the promising antioxidant action of Lamiaceae herbs in vitro, human studies on these potential sources of dietary antioxidants have remained scarce. In this work, the phenolic acids recovered in human urine after single ingestion of Origanum onites extract were analyzed. The excretion was increased 4- and 2-fold during 0-24 and 24-48 h of the follow-up, respectively. The mean increase in the excretion of phenolic compounds exceeded the ingested amount of identified phenolic acids. The result can be partly explained by rosmarinic acid, the main identified phenolic constituent in the extract, as well as flavonoids present in minor amounts, presumably being metabolized into a double amount of simple phenolic metabolites. Furthermore, unidentified phenolic constituents in the extract partly contribute to the excretory increase. The main metabolite, p-hydroxybenzoic acid, was excreted rapidly. The results show that constituents of oregano extract and, in particular, their metabolites may contribute to the dietary intake of phenolic antioxidants.

Fractionation of polyphenols in hawthorn into polymeric procyanidins, phenolic acids and flavonoids prior to high-performance liquid chromatographic analysis.

Polymeric procyanidins, phenolic carboxylic acids and flavonoids of hawthorn (Crataegus laevigata) were fractionated prior to HPLC analysis using column chromatography and solid-phase extraction (SPE). The flavonoid fraction also contained (-)-epicatechin. The three groups of phenolics, each with clearly different UV spectra, were examined by means of high-performance liquid chromatography-diode array detection (HPLC-DAD) analysis. The average repeatability of the method (RSD) was in the range of 8-13% for chlorogenic acid, (-)-epicatechin and hyperoside. The polymeric procyanidins of hawthorn flowers consisted mainly of (-)-epicatechin subunits, and their mean degree of polymerization (DP) was 22.2. The HPLC methods developed can be used for the qualitative and quantitative analysis of different phenolic compounds in hawthorn plant material and their extracts.

Chemical composition and in vitro antioxidant evaluation of commercial water-soluble willow herb (Epilobium angustifolium L.) extracts.

Five commercially available water-soluble extracts prepared from the aerial parts of Epilobium angustifolium L. (Onagraceae) were screened for antioxidant-related properties in a battery of six in vitro assays. Total phenol content and qualitative-quantitative analyses were also carried out. The extracts demonstrated varying degrees of efficacy in each screen. Two extracts, denoted as nonfermented and Tver, were the most effective toward reducing iron(III), scavenging 1,1-diphenyl-2-picrylhydrazyl free radicals, inhibiting hydroxyl radical-catalyzed bovine brain-derived phospholipid degradation, and non-site- and site-specific hydroxyl radical-catalyzed 2-deoxy-D-ribose degradation. The activity profile of the extracts changed, however, when their iron(II) chelating ability was assessed. The nonfermented and Tver extracts were not as effective iron(II) chelators as the extract denoted as Lotos. All the extracts contained Folin-Ciocalteu-reactive substances, which was confirmed by the presence of predominantly polar phenolic analytes (i.e., hydroxylated benzoic acid derivatives and flavonoids).