Damiano Rossi

Bioactivities of Piper aduncum L. and Piper obliquum Ruiz & Pavon (Piperaceae) essential oils from Eastern Ecuador

Essential oils from aerial parts of Piper aduncum (Matico) and Piper obliquum (Anis del Oriente) of ecuadorian origin were analyzed by GC-FID, GC-MS, (13)C NMR and their biological and pharmacological activities were assessed. Chemical composition proved to be unusually different from previous reports for safrole-rich P. obliquum (45.8%), while P. aduncum main constituent was dillapiol (45.9%). No genotoxic activity was found in the Ames/Salmonella typhimurium (TA98 and TA100) assay, either with or without S9 activation. Mutagen-protective properties, evaluated using sodium azide, 2-nitrofluorene and 2-aminoanthracene as mutagens/promutagens, was observed against promutagen 2-aminoanthracene, likely in consequence of microsomial deactivation. Antimicrobial assays have been performed on Gram+/Gram- bacteria, dermatophyte and phytopathogenic fungi and best results were provided by P. aduncum against fungal strains with complete inhibition at 500μg/ml. Preliminary analgesic and antithrombotic activities evidenced the absence of the former in hot plate and edema assays and a limited antiplatelet action against three different agonists (ADP, AA and U46619). Both oils have a very limited antioxidant capacity.

Clotrimazole nanoparticle gel for mucosal administration.

In this study a formulation suitable to be applied on oral and/or vaginal mucosa has been developed for the treatment of fungal infections. The aim of the research is a comparison between clotrimazole (CLO) containing semisolid formulations based on monoolein aqueous dispersion (MAD) or nanostructured lipid carrier (NLC). MAD and NLC have been characterized in terms of morphology and dimensional distribution by cryogenic Transmission Electron Microscopy (cryo-TEM) and Photon Correlation Spectroscopy (PCS). CLO was encapsulated with high entrapment efficiency both in MAD and in NLC, according to Sedimentation Field Flow Fractionation (SdFFF) combined with HPLC. CLO recovery in MAD and NLC has been investigated by time. In order to obtain formulations with suitable viscosity for mucosal application, MAD was diluted with a carbomer gel, while NLC was directly viscosized by the addition of poloxamer 407 in the dispersion. The rheological properties of MAD and NLC after viscosizing have been investigated. Franz cell has been employed to study CLO diffusion from the different vehicles, evidencing diffusion rates from MAD and NLC superimposable to that obtained using Canesten(®). An anticandidal activity study demonstrated that both CLO-MAD and CLO-NLC were more active against Candida albicans with respect to the pure drug.

trans-Resveratrol in Nutraceuticals: Issues in Retail Quality and Effectiveness

Fourteen brands of resveratrol-containing nutraceuticals were evaluated in order to verify their actual resveratrol content and to control if their health-promoting properties are related to manufacturing quality. Products included pure resveratrol capsules or multi-ingredient formulations with standardized amounts of resveratrol and other phytochemicals. Samples were analyzed for total trans-resveratrol, flavonoids, procyanidin, polyphenol content and the results were compared with the content declared on-label. Only five out of 14 brands had near label values, compliant with Good Manufacturing Practices (GMP) requirements (95–105% content of active constituent), four products were slightly out of this range (83–111%) and three were in the 8–64% range. Two samples were below the limit of detection. The greater the difference between actual and labeled resveratrol content, the lower was the antioxidant and antiproliferative activity strength. Dietary supplements containing pure trans-resveratrol exhibited a greater induction of differentiation towards human leukemic K562 cells when compared to multicomponent products. Great differences currently exist among resveratrol food supplements commercially available and GMP-grade quality should not be taken for granted. On the other side, dosages suggested by most “pure”, “high-dosage” supplements may allow a supplementation level adequate to obtain some of the purported health benefits.

Effect of high hydrostatic pressure and high pressure homogenization on the enantioselectivity of microbial reductions

Abstract The effect of high hydrostatic pressure and high pressure homogenization on the microbial reductions of the model prochiral ketones 1a-c is described. Various strains of Saccharomyces cerevisiae and Yarrowia lipolytica are utilized in the reduction and higher enantioselectivities are generally achieved together with lower yields compared with the results obtained at atmospheric pressure. In some cases both increasing yields and inversion of enantioselectivity are reported. The effects of the hydrostatic and the homogenization pressure are also compared with the cell viability.

Metabolite profiling of polyphenols in a Terminalia chebula Retzius ayurvedic decoction and evaluation of its chemopreventive activity.

ETHNOPHARMACOLOGICAL RELEVANCE The decoction of Terminalia chebula fruit is an ayurvedic remedy whose prolonged oral administration is prized as a generic intestinal and hepatic detoxifying agent. Its administration is suggested also under the perspective of a reduced risk of cancer, metabolic and cardiovascular diseases. AIM OF THE STUDY To evaluate the phytochemical profile and the chemopreventive potential of Terminalia chebula fruit decoction prepared according to the ayurvedic decoction recipe. MATERIALS AND METHODS The quali- and quantitative metabolite profiling of polyphenols was obtained using HPLC-UV/DAD and HPLC-ESI-MS. The crude decoction and purified compounds were tested for their capability to interact with the EphA2-ephrin-A1 system and for their antimutagenic properties against dietary and environmental mutagens (AA, 2-NF, NaN3, and heterocyclic amines IQ, MeIQ, MeIQx, Glu-P1, Glu-P2,) in the Ames-Salmonella/microsome assay, with and without enzymatic induction. RESULTS The decoction was found to contain 3,4,6-tri-O-galloyl-d-glucose (55.87 mg/g), chebulic acid (54.03 mg/g), β-punicalagin (41.25mg/g), corilagin (40.31 mg/g), α-punicalagin (35.55 mg/g), chebulagic acid (29.09 mg/g), gallic acid (27.96 mg/g) 1,3,4,6-tri-O-galloyl-β-d-glucose (24.25mg/g) chebulinic acid (20.23 mg/g), 1,2,3,4,6-penta-O-galloyl-d-glucose (13.53 mg/g), ellagic acid (8.00 mg/g), 1,6-di-O-galloyl-d-glucose (4.16 mg/g). An inhibitory effect was recorded in both Salmonella typhimurium TA98 and TA100 strains against the mutagenic activity of heterocyclic amines (22-61%), promutagen AA (91-97%) and directly acting mutagen 2-NF (52%) with but not against NaN3 (7%). Galloyl derivatives allowed an inhibition of mutagenicity induced by MeIQ above 80% at 0.01 mol/plate. Both decoction and purified compounds were able to modulate the EphA2-ephrinA1 system, suggesting a potential multiple chemopreventive mechanism. CONCLUSIONS The traditional ayurvedic decoction of Terminalia chebula may harbour a potential as a safe and low-cost chemopreventive agent at the intestinal level, if administered according to the ayurvedic specifications. Moreover, its recourse may enhance the presence of some polyphenolic constituents.

Bergamot (Citrus bergamia Risso) fruit extracts and identified components alter expression of interleukin 8 gene in cystic fibrosis bronchial epithelial cell lines

BackgroundCystic fibrosis (CF) airway pathology is a fatal, autosomal, recessive genetic disease characterized by extensive lung inflammation. After induction by TNF-α, elevated concentrations of several pro-inflammatory cytokines (i.e. IL-6, IL-1β) and chemokines (i.e. IL-8) are released from airway epithelial cells. In order to reduce the excessive inflammatory response in the airways of CF patients, new therapies have been developed and in this respect, medicinal plant extracts have been studied. In this article we have investigated the possible use of bergamot extracts (Citrus bergamia Risso) and their identified components to alter the expression of IL-8 associated with the cystic fibrosis airway pathology.MethodsThe extracts were chemically characterized by 1H-NMR (nuclear magnetic resonance), GC-FID (gas chromatography-flame ionization detector), GC-MS (gas chromatography-mass spectrometry) and HPLC (high pressure liquid chromatography). Both bergamot extracts and main detected chemical constituents were assayed for their biological activity measuring (a) cytokines and chemokines in culture supernatants released from cystic fibrosis IB3-1 cells treated with TNF-α by Bio-Plex cytokine assay; (b) accumulation of IL-8 mRNA by real-time PCR.ResultsThe extracts obtained from bergamot (Citrus bergamia Risso) epicarps contain components displaying an inhibitory activity on IL-8. Particularly, the most active molecules were bergapten and citropten. These effects have been confirmed by analyzing mRNA levels and protein release in the CF cellular models IB3-1 and CuFi-1 induced with TNF-α or exposed to heat-inactivated Pseudomonas aeruginosa.ConclusionsThese obtained results clearly indicate that bergapten and citropten are strong inhibitors of IL-8 expression and could be proposed for further studies to verify possible anti-inflammatory properties to reduce lung inflammation in CF patients.

Bergamot (Citrus bergamia Risso) Fruit Extracts as γ-Globin Gene Expression Inducers: Phytochemical and Functional Perspectives

Epicarps of Citrus bergamia fruits from organic farming were extracted with the objective of obtaining derived products differently rich in coumarins and psoralens. The extracts were chemically characterized by (1)H nuclear magnetic resonance (NMR), gas chromatography-flame ionization detection (GC-FID), gas chromatography-mass spectrometry (GC-MS), and high-pressure liquid chromatography (HPLC) for detecting and quantifying the main constituents. Both bergamot extracts and chemical standards corresponding to the main constituents detected were then assayed for their capacity to increase erythroid differentiation of K562 cells and expression of γ-globin genes in human erythroid precursor cells. Three experimental cell systems were employed: (a) the human leukemic K562 cell line, (b) K562 cell clones stably transfected with a pCCL construct carrying green-enhanced green fluorescence protein (EGFP) under the γ-globin gene promoter, and (c) the two-phase liquid culture of human erythroid progenitors isolated from healthy donors. The results suggest that citropten and bergapten are powerful inducers of differentiation and γ-globin gene expression in human erythroid cells. These data could have practical relevance, because pharmacologically mediated regulation of human γ-globin gene expression, with the consequent induction of fetal hemoglobin, is considered to be a potential therapeutic approach in hematological disorders, including β-thalassemia and sickle cell anemia.

Neuroprotective and anti-inflammatory properties of a novel non-thiazolidinedione PPARγ agonist in vitro and in MPTP-treated mice.

Peroxisome proliferator-activated receptor (PPAR)γ is a potential pharmacological target for disease-modification in Parkinsons disease (PD), mainly acting by modulating the neuroinflammatory response. However, currently available agonists thiazolidinediones (TZDs) present limitations due to safety concerns. We evaluated a novel thiobarbituric-like compound MDG548, which acts as a functional PPARγ agonist displaying higher and selective binding affinity as compared to TZDs. Neuroprotection by MDG548 was tested in vitro and in a mouse MPTP model of PD, and neuroinflammation was investigated as a putative underlying mechanism. Viability assay on rat cortical neurons showed lack of cytotoxic effect in the dose-range of 100 nM-10 μM, which was therefore used for testing in vitro protection against H2O2 and MPP+ neurotoxicity. MDG548 dose-dependently increased cell viability of rat cortical neurons co-treated with H2O2 or pre-exposed to MDG548 prior to H2O2. Moreover, MDG548 induced neuroprotection in MPP+-treated PC12 cells. NF-kB activation was investigated to assess anti-inflammatory activity. MDG548 dose-dependently decreased NF-kB activation induced by LPS (100 ng/100ml) in HEK-Blue-hTLR4 cells. Given the supposed cancer risk of other PPARγ agonists, Ames test for genotoxicity was performed in Salmonella typhimurium TA100 and TA98 strains, showing that MDG548 was not genotoxic. In vivo, BL/6J mice were treated with MPTP (20mg/kg i.p. once/day for 4 days) in association with saline or MDG548 (2, 5, 10 mg/kg i.p.). Stereological counting showed that MDG548 prevented the MPTP-induced reduction in TH-positive cells in the substantia nigra compacta (SNc) at all doses tested. Moreover, MDG548 reduced reactive microglia and iNOS induction in the SNc. MDG548, being a non-TZD compound with high PPARγ affinity, void of genotoxicity, and with in vitro as well as in vivo neuroprotective properties, provides a promising alternative in the search for safer PPARγ agonists to be tested as potential disease-modifying drugs in PD.

Metabolite profiling of polyphenols in Vaccinium berries and determination of their chemopreventive properties

A detailed investigation on the chemical composition and chemopreventive activity of Vaccinium floribundum Kunth berries was carried out in comparison with Vaccinium myrtillus L. Berry polyphenols were extracted by using two sequential dynamic maceration steps, which enabled to maximize the yields of secondary metabolites. In particular, phenolic acids and flavonols were extracted from berries using ethyl acetate (EtOAc), whereas anthocyanins were extracted from the residue with 0.6M HCl in methanol (MeOH). The analysis of secondary metabolites in berry extracts was performed by means of two specific HPLC methods. Phenolic acids and flavonols were analyzed on an Ascentis C18 column (250mm×4.6mm I.D., 5μm), with a gradient mobile phase composed of 0.1M HCOOH in H2O and ACN. Anthocyanin analysis was carried out on a Zorbax SB-C18 column (150mm×4.6mm I.D., 5μm), with a gradient mobile phase composed of H2O-HCOOH (9:1, v/v) and MeOH-H2O-HCOOH (5:4:1, v/v/v). Detection was performed by UV/DAD, MS and MS(2). The polyphenol composition of V. floribundum and V. myrtillus was studied in detail. The samples of V. floribundum analyzed in this study had a much higher content of both phenolic acids and flavonols in comparison with V. myrtillus (mean value 41.6±10.2 and 13.7±0.2mg/100g FW, respectively), while V. myrtillus showed a higher amount of anthocyanins if compared with V. floribundum (568.8±8.8 and mean value 376.2±49.9mg/100gFW, respectively). The extracts gave negative results in antimutagenic assays against carcinogens 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ) and 4-nitroquinoline-1-oxide (4-NQO), while they performed similarly in both ABTS(+) and DPPH antioxidant assays.

Polyphenol rich botanicals used as food supplements interfere with EphA2-ephrinA1 system

The Eph tyrosine kinase receptors and their ephrin ligands play a central role in several human cancers and their deregulated expression or function promotes tumorigenesis, inducing aggressive tumor phenotypes. Green tea extracts (GTE) have been recently found to inhibit Eph-kinase phosphorylation. In order to evaluate the potential contribution of edible and medicinal plants on EphA2-ephrinA1 modulation, 133 commercially available plant extracts used as food supplements, essential and fixed oils were screened with an ELISA-based binding assay. Nine plant extracts, rich of polyphenols, reversibly inhibited binding in a dose-dependent manner (IC₅₀ 0.83-24 μg/ml). Functional studies on PC3 prostate adenocarcinoma cells revealed that active extracts antagonized ephrinA1-Fc-induced EphA2-phosphorylation at non-cytotoxic concentrations (IC₅₀ 0.31-11.3 μg/ml) without interfering with EGF-induced EGFR activation, suggesting a specific effect. These findings could furnish an interesting starting point regarding the potential relationship between diet, edible plant secondary metabolites and Eph-ephrin system, suggesting their possible involvement in cancer development modulation.