OBJECTIVE:Fluxonorm® is a dietary supplement that includes water-soluble extracts of Solidago virga-aurea, Phyllantus niruri, Epilobium angustifolium, Peumus boldus and Ononis spinosa. The aim of the present study was to evaluate the tolerability and efficacy of Fluxonorm® in improving lower urinary tract symptoms in patients with benign prostatic hyperplasia (BPH) in combination with standard of care.PATIENTS AND METHODS:Lower urinary tract symptoms can be improved by a marked anti-inflammatory action on the lower urinary tract (irritative symptoms) and/or by an anti-proliferative action (obstructive symptoms) on the prostate. Thirty patients were enrolled to evaluate the effect of Fluxonorm® on improving lower urinary tract symptoms. All patients complained of lower urinary tract symptoms (LUTS), such as hesitancy, poor flow, intermittent flow, incomplete voiding (obstructive symptoms), as well as increased frequency, nocturia and urgency (storage symptoms). All patients were treated with one tablet of Fluxonorm® (1200 mg) daily for 30 days to corroborate the results of our observation in which the food supplement (800 µg/mL) was also studied on the human prostate cancer PC3 cell line (antiproliferative activity) and on prostaglandin (PG)E2 production (anti-inflammatory activity). In addition, the effect of this compound on cyclooxygenase-2 (COX-2) gene expression was investigated. Finally, a bioinformatic analysis was conducted with the aim of unravelling the mechanism of action underlying the observed bio-pharmacological effects.RESULTS:As hypothesized in our preclinical research, adding Fluxonorm® to the therapy of enrolled patients improved all studied clinical parameters, including maximum flow (Qmax), after one month of treatment. In the preclinical evaluation, this formulation reduced PC3 cell viability and PGE2 production. The effects were also paralleled by reduced COX-2 gene expression and Fluxonorm®'s partly related content of catechin. While docking studies pointed out to the putative inhibition of matrix metalloproteinse-2 by gallic acid, as a further mechanism underlying the observed anti-proliferative effects, in PC3 cells exposed to Fluxonorm®.CONCLUSIONS:Fluxonorm® improved the efficacy of standard therapy, in terms of antioxidant/anti-inflammatory effects, for the management of lower urinary tract symptoms (LUTS). This could be related, albeit partially, to the blunting effect of this compound on PGE2 production.

Ethyl acetate (EA), methanol (MeOH), and aqueous extracts of aerial parts of Anthemis tinctoria var. pallida (ATP) and A. cretica subsp. tenuiloba (ACT) were investigated for their phenol and flavonoid content, antioxidant, and key enzyme inhibitory potentials. All extracts displayed antiradical effects, with MeOH and aqueous extracts being a superior source of antioxidants. On the other hand, EA and MeOH extracts were potent against AChE and BChE. Enzyme inhibitory effects against tyrosinase and α-glucosidase were observed, as well. We also studied Anthemis extracts in an ex vivo experimental neurotoxicity paradigm. We assayed extract influence on oxidative stress and neurotransmission biomarkers, including lactate dehydrogenase (LDH) and serotonin (5-HT), in isolated rat cortex challenged with K+ 60 mM Krebs-Ringer buffer (excitotoxicity stimulus). An untargeted proteomic analysis was finally performed in order to explore the putative mechanism in the brain. The pharmacological study highlighted the capability of ACT water extract to blunt K+ 60 mM increase in LDH level and 5-HT turnover, and restore physiological activity of specific proteins involved in neuron morphology and neurotransmission, including NEFMs, VAMP-2, and PKCγ, thus further supporting the neuroprotective role of ACT water extract.

Multiple studies revealed the potential application of high quality saffron byproducts as cheap sources of bioactive compounds endowed with antioxidant activity. In the present study, we analyzed the total fatty acids of the anthers, and explored the pharmacological and toxicological potential of anthers, by evaluating genotoxic and protective effects in multiple cell lines, brine shrimps and isolated rat tissues.

Origanum sipyleum L., an endemic plant of Western Anatolia has been used as a medicinal tea, food additive, and for the production of essential oil. In this study, the biological potential of three extracts (ethyl acetate, methanol, and aqueous) of O. sipyleum was assessed based on antioxidant activity against key enzymes of clinical relvance. The chemical profile of the plant was assessed using spectrophotometric and LC-MS techniques. Additionally, we explored potential antioxidant and anti-inflammatory effects duced by the extracts in an experimental model of ulcerative colitis induced by LPS challenging. LC-MS analysis revealed that the extracts contained different classes of phenolics, such as rosmarinic acid, phlorizin and gallic acid. We found that the aqueous extract was the most effective antioxidant, displaying the highest DPPH and ABTS scavenging, FRAP, CUPRAC, molybdenum(VI) reducing, and metal chelating effect. The aqueous extract howed the strongest acetylcholinesterase (AChE) inhibition; the methanol extract showed the highest α-glucosidase inhibition, while the ethyl acetate extract was the most effective on butyrylcholinesterase (BChE), tyrosinase, and α-amylase. The total flavonoid content was highest in the aqueous and ethyl acetate extract, respectively. Finally, we found that all extracts were effective in reducing LPS-induced activity of pro-oxidant and pro-inflammatory biomarkers including nitrites, LDH, PGE2 and 5-HT, in rat colon, with the best activity showed by ethyl acetate extract. Our results indicated that the three solvent extracts varied in their chemical and biological profiles, but overall, O. sipyleum showed promising therapeutic properties, nonetheless, need to be further validated in in vivo models.

One of the most promising economic perspectives of hemp production chain is female inflorescence valorization. By contrast, scientific literature lacks on chemical composition or biological activity data from aqueous fraction obtained from industrial hemp flowers, which have long been considered as waste products. In this context, the main focus of the following study is the evaluation of antioxidant and anti-inflammatory effects related to aqueous flower extracts from four commercial hemp cultivars (Futura 75, Kc virtus, Carmagnola Cs and Villanova). We evaluated the extract phytochemical profile. Then, we studied the water extracts both in vitro and ex vivo in order to test protective effects in an experimental model of ulcerative colitis, constituted by isolated LPS-stimulated colon. All cultivar extracts displayed similar total phenol and flavonoid content. On the other hand, Futura 75 cultivar extract displayed a better antioxidant and anti-inflammatory profile. Considering this, Futura 75 extract has been subsequently tested to evaluate its effect on pathogen bacterial and fungal species involved in ulcerative colitis, finding a significant inhibition on the growth of C. albicans and selected Gram positive and negative bacterial strains.

Prostatitis is a common prostate disease that could be promoted by bacterial or non-bacterial infectious agents. In addition, inflammatory pathways involved in prostatitis have been increasingly studied, and herbal extracts endowed with anti-inflammatory effects are under investigation, individually or in combination, for their efficacy in alleviating the burden of inflammation, with possible improvements in symptoms. Serenoa repens (Serenoa), in combination with Crocus sativus (Crocus) and Pinus massoniana (Pinus), has previously shown to improve sexual function and limit urinary symptoms in patients suffering from concomitant erectile dysfunction and lower urinary tract symptoms. In this context, the aim of the present study is to evaluate the efficacy of Serenoa, Crocus and Pinus extracts, either alone or in combination, on immortalized prostate cells (PC3) and in an experimental model of bacterial prostatitis constituted by ex vivo prostate specimens challenged with lipopolysaccharide (LPS). We found that the tested extracts were able to reduce ROS production by PC3 cells and NFkB and PGE2 activity in prostate specimens challenged with LPS. In addition, the pharmacological association of the extracts displayed synergistic effects indicating a rational use of the mixture of the tested extracts as a novel anti-oxidant and anti-inflammatory formulation in bacterial prostatitis. Finally, we performed analytical and in vitro evaluation to better characterize the phytochemical profile and the mechanism of action of selected secondary metabolites.

Resveratrol (3,4',5-trihydroxy-trans-stilbene) is a natural phytoalexin found in grapes and wine, which has been extensively studied for a wide range of biological effects. A large number of stilbene-containing derivatives have displayed antioxidant and antiproliferative activities on various cancer cell lines. In this study, a series of stilbene hybrids 1-9, previously reported as peroxisome proliferator-activated receptor (PPAR) agonists, were assessed at micromolar concentrations using MTT cell viability assay in C2C12 and MCF7 cell lines. The modulation of oxidative stress was also evaluated by measuring the protective effects on reactive oxygen species (ROS) production induced or not by oxidative stimulus. Among these, compounds 2 and 8 showed significant radical scavenging activity.

Copyright © by BIOLIFE, s.a.s. This publication and/or article is for individual use only and may not be further reproduced without written permission from the copyright holder. Unauthorized reproduction may result in financial and other penalties DISCLOSURE: ALL AUTHORS REPORT NO CONFLICTS OF INTEREST RELEVANT TO THIS ARTICLE. JOURNAL OF BIOLOGICAL REGULATORS & HOMEOSTATIC AGENTS Vol. 31, no. 3, 531-541 (2017)

A pivotal role in osteoporosis development is played by radical oxygen species (ROS), the increased production of which is related to inhibited osteoblastic activity and bone formation. A new field of research could involve medicinal plants with antioxidant and protective effects in osteoporosis. Furthermore, considering the multifactorial metabolic aspects of osteoporosis, the pharmacological association of multiple medicinal plants could improve patient response. The aim of the present study is to evaluate in vitro and in vivo the protective effects of a natural formula containing lactoferrin 12%, Equisetum arvensis ES 54%, soy isoflavones 34% and vitamin D3 0.002%, in PBMC and C2C12 cells and in the bone matrix of young (3-month-old) and aged (12-month-old) female Sprague-Dawley rats, following chronic (21 days) administration. In this context, we assayed the activities of several inflammation and bone homeostasis mediators, such as IL-6, TNFα, PGE2, osteoprotegerin, RANK, RANKL and NFkB. In vitro studies showed that natural formula (5-1000μg/ml) was able to significantly inhibit ROS and PGE2 production. In the same concentration range, the natural formula inhibited both TNFα and IL-6 gene expression. In the in vivo studies, we administered to young and aged female rats the natural formula at 5mg/rat for 21 days, finding a significant reduction in inflammatory PGE2 and NFkB activity. Nevertheless, we observed a significant increase in osteoprotegerin/RANKL ratio only in aged rats, compared to the respective control group. In conclusion, our findings corroborate the rational use of natural formula in the prevention and management of osteoporotic disease.

This study investigated the effects of rosemary dietary supplementation on the rumen degradability of alfalfa hay, the ruminal bacterial population and rumen metabolism in sheep. The experiment was designed as a 4×4 Latin square balanced for carry-over effects using four ruminally cannulated sheep fed a basal diet consisting of 1.5kg/day of alfalfa hay supplemented (400g/day) with one of four different concentrates as follows: (1) control (CTR); (2) CTR with the addition of 1.75% rosemary essential oil adsorbed on an inert support (EO); (3) CTR with the addition of 2.50% dried and ground rosemary leaves (RL); and (4) same as (3), except that rosemary leaves were included in the concentrate before pelleting (RL pellet). No effects were recorded in the volatile fatty acid or lactic acid production. Ruminal ammonia concentration tended to be lower (P=0.077) with the RL pellet diet. The crude protein degradability, after incubation times of 2 and 48h, decreased (P<0.001) in sheep fed with the RL pellet diet. The dry matter degradability followed the same trend, with lower (P<0.001) rates of disappearance resulting from the RL pellet diet after 24h of incubation. The pelleting process may influence the availability of the active compounds of rosemary leaves, allowing greater antibacterial activity. Supplementation of sheep diets with rosemary leaves could potentially be used to manipulate ruminal degradation patterns.

Imoviral™ is a natural product formulation containing a mixture of uncaria, shiitake and ribes extracts. All ingredients are recognized as antioxidant, anti-inflammatory agent and immunomodulant. In order to evaluate the rational basis of extract mixture as immunomodulatory agent, we tested the effect of Imoviral™ formulation on macrophage response to lipopolysaccharide (LPS)-induced stress. The effect was evaluated as variation of reactive oxygen species (ROS) and prostaglandin E2 (PGE2) production and as cytokine gene expression. The extract did not affect cell viability up to 250 μg/ml. Treatment with extract (10-150 μg/ml) reduced ROS and PGE2 production as well as IL-8 and TNF-α gene expression. A pre-treatment with extract blunted LPS-induced production of ROS and PGE2, markers of oxidative and inflammatory stress, as well as the gene expression of all cytokines tested, indicators, in vitro, of immune response activation. In conclusion, we demonstrated that Imoviral™ formulation could be a useful tool to modulate the immune function, reducing the oxidative and inflammatory markers related to bacterial attack. Experimental data suggest that Imoviral™ extract mixture could also represent a preventive pharmacological strategy to enhance cell resistance to bacterial infections.

AIM:Aim of this study was to evaluate the effects of phytocomplexes of Uncaria, Shiitake and Ribes in terms of viability and inflammatory response on immune cell-derived cultures.METHODS:Standardized extracts of Uncaria, Shitake and Ribes and their commercial formulation were tested on cell lines PBMC, U937 and macrophage. The activity was evaluated in terms of cell viability (MTT test), variations of oxidative marker release (ROS and PGE2) and modulatory effects on immune response (gene expression of IL-6, IL-8 and TNFα, RT-PCR).RESULTS:Cell viability was not affected by extracts, except subtle variations observed only at higher doses (>250 µg/mL). The extract mixture was well tolerated, with no effects on cell viability up to doses of 500 µg/mL. Pre-treatment of macrophages with subtoxic doses of the extracts reduced the basal release of oxidative markers and enhanced the cell response to exogenous oxidant stimulation, as revealed by ROS and PGE2 release reduction. The same treatment on macrophage resulted in a selective modulation of the immune response, as shown by an increase of IL-6 mRNA and, partially, IL-8 mRNA, while a reduction was observed for TNFα mRNA.CONCLUSION:Data confirm that extracts and their formulations can act as regulator of the immune system with mechanisms involving the oxidative stress and the release of selected proinflammatory cytokines.

Aim. Aim of this study was to evaluate the effects of phytocomplexes of Uncaria, Shitake and Ribes in terms of viability and inflammatory response on immune cell-derived cultures. Methods. Standardized extracts of Uncaria, Shitake and Ribes and their commercial formulation were tested on cell lines PBMC, U937 and macrophage. The activity was evaluated in terms of cell viability (MTT test), variations of oxidative markers release (ROS and PGE2) and modulatory effects on immune response (RT-PCR expression of IL-6, IL-8 and TNFα). Results. Cell viability was not affected by extracts, except subtle variations observed only at higher doses (>250 μg/mL). The extract mixture was well tolerated, with no effects on cell viability up to doses of 500 μg/mL. Pre-treatment of macrophages with subtoxic doses of the extracts reduced the basal release of oxidative markers and enhanced the cell response to oxogenus oxidant stimulation, as revealed by ROS and PGE2 release reduction. The same treatment on macrophage resulted in a selective modulation of the immune response, as shown by an increase of IL-6 mRNA and, partially, IL-8 mRNA, while a reduction was observed for TNFα mRNA. Conclusion. Data confirm that extracts and their formulations can act as regulator of the immune system with mechanisms involving the oxidative stress and the release of selected proinflammatory cytokines.

Plants of cranberry (Vaccinium macrocarpon) furnish edible fruits and derivates that have been used for the prevention and treatment of urinary tract infections. In the present work we compare two commercial extracts that contain proanthocyanins (PACs) at 4 percent and 20 percent for antimicrobial, antiproliferative, antiradical and protective properties against oxidative stress on cell lines. Both extracts showed antimicrobial activity (MIC values range 3-100 microg/ml). Extract at 20 percent PACs showed higher antiproliferative activity against HepG2 and MCF7 cells, but not against C2C12 cells. Both extracts showed a dose-dependent free-radical scavenging capacity, and a protective effect on the cell damage was also revealed by reduction of intracellular active oxygen species release. Cranberry extracts confirmed antioxidative properties and efficacy in reduction of cell viability that resulted stronger against tumor cells. The pretreatment with cranberry extracts, furthermore, reveal an increase of cell resistance against oxidative stress, suggesting a potential role as a dietary supplement in preventing free-radical damage. The proanthocyanidin content is critical to determine the extract efficacy. In cellular experiments the extracts resulted clearly differentiated in their activity, and the activity was strongly influenced by PACs content. Only in DPPH test the free radical scavenging activity seemed to be directly related to proanthocyanidins content.

INTRODUCTION Rhamnus alpinus L. (Rhamnaceae), a traditional plants in the flora of the Abruzzo region, is known to contain active anthraquinone secondary metabolites. However, the content of anthraquinones varies among R. alpinus samples depending on collection season and site. Thus, using simple, reliable and accurate analytical methods for the determination of anthraquinones in R. alpinus extracts allows comparative study of different methods of extraction. OBJECTIVE After a partial validation of an HPLC method for the simultaneous determination of five anthraquinones, aloe-emodine, rheine, emodine, chrysophanol and physcione, in the bark of R. alpinus, we compared three different methods of extraction. METHODOLOGY Anthraquinones were extracted from the bark of R. alpinus using different techniques (methanol maceration, ultrasonic and supercritical CO(2) extraction). Separation and quantification of anthraquinones were accomplished using a reversed-phase C(18) column with the mobile phase of H(2)O-methanol (40 : 60, v/v, 1% formic acid) at a wavelength of 254 nm. The qualitative analyses were also achieved at wavelength of 435 nm. RESULTS All calibration curves were linear over the concentration range tested (10-200 mM) with the determination coefficients >or=0.991. The detection limits (S/N = 3) were 5 mM for each analytes. All five anthraquinones were found in the samples tested at concentrations reported in experimental data. CONCLUSION The described HPLC method and optimised extraction procedure are simple, accurate and selective for separation and quantification of anthraquinones in the bark of R. alpinus and allow evaluation of the best extraction procedure between the tested assays.

Artichoke, dandelion, turmeric extracts and rosemary essential oil are commonly used as ingredients in many herbal preparations to treat hepatic and gallbladder disorders. In the present work we compare the activity of each single extract with a commercial mixture for antiproliferative, antiradical and protective effects against induced oxidant stress effect. In ABTS and DPPH tests, turmeric extract is the most active, followed by artichoke and dandelion. All samples exhibited antiproliferative activity in a dose-dependent manner against HepG2 cells. In the same cell lines, the protective effect of pre-treatment with the extracts were detected by evaluating the prostaglandin E2 release, a marker of oxidative stress induced by hydrogen peroxide. The treatments with the extracts were efficient in reducing the release of PGE2 induced by oxidative stimulus. The positive results of the cell viability test, together with the protective and antiradical activity confirm the rationale for the use of these ingredients in commercial formulations as a health aid tool in modern phytotherapy.

Several species belonging to the genus Rhamnus (Rhamnaceae), comprising ones among which are found the most typical plants of the Italian flora, are known to contain biologically active anthraquinone secondary metabolites. Although several Rhamnus species were so far investigated, no information is available concerning the content and relative abundances of anthraquinones in R. saxatilis. In this study we used a simple, reliable, and accurate analytical method to determine the anthraquinones in bark of R. saxatilis. This allowed us also to trace a comparative study on the efficacy of different extraction solvents in ultrasonication time dependent assays. Separation and quantification of anthraquinones were accomplished using a C18 column with the mobile phase of H2O:methanol (40:60, v/v, 1% formic acid) at a flow rate of 0.7mL/min and a detection wavelength of 254nm, while the qualitative analyses were also achieved at a wavelength of 435nm.Finally, the described HPLC method, was used to obtain a specific chemical fingerprint for this species in comparison with other species from the same family.