Bioactive food components for colorectal cancer prevention and treatment: A good match.

Colorectal cancer (CRC) is the third most frequent cancer worldwide, accounts for about 10% of the total cancer cases, and ranks as the second cause of death by cancer. CRC is more prevalent in developed countries in close causal relation with occidental diets. Due to anatomy, the diet has a strong impact on CRC. High contents in meat are acknowledged risk factors whereas a diet rich in fruits and vegetables is an established CRC protective factor. Fruits and vegetables contain numerous Bioactive Food Components (BFCs), physiologically active food compounds, beneficial on health. Preventive and therapeutic benefits of BFCs in cancer have increasingly been reported over the past 20 years. BFCs show both chemopreventive and anti-tumor properties in CRC but more interestingly, abundant research describes BFCs as enhancers of conventional cancer treatments. Despite these promising results, their clinical transferability is slowed down by bioavailability interrogations and their poorly understood hormetic effect. In this review, we would like to reposition BFCs as well-fitted for applications in CRC. We provide a synthetic overview of trustworthy BFC applications in CRC, with a special highlight on combinatory approaches and conventional cancer treatment potentiation strategies.

Transport of hydrocortisone in targeted layers of the skin by multi-lamellar liposomes.

Hydrocortisone (HyC), a hydrophobic pharmaceutical active, was encapsulated in multi-lamellar liposomes (MLLs) composed of P100, a mixture of phospholipids, and Tween®80. Three different HyC-loaded formulations were designed to target the stratum corneum, the living epidermis and the hypodermis. The impact of encapsulation on their size, elasticity and zeta potential, the three key factors controlling MLLs skin penetration, was studied. Raman mapping of phospholipids and HyC allowed the localisation of both components inside an artificial skin, Strat-M®, demonstrating the efficiency of the targeting. Percutaneous permeation profiles through excised human skin were performed over 48 h, supporting results on artificial skin. Their modelling revealed that HyC encapsulated in MLLs, designed to target the stratum corneum and living epidermis, exhibited a non-Fickian diffusion process. In contrast, a Fickian diffusion was found for HyC administered in solution, in a pharmaceutical cream and in transdermal MLLs. These results allowed us to propose a mechanism of interaction between HyC-containing MLLs and the skin.

Resveratrol, ε-Viniferin, and Vitisin B from Vine: Comparison of Their In Vitro Antioxidant Activities and Study of Their Interactions.

The control of oxidative stress with natural active substances could limit the development of numerous pathologies. Our objective was to study the antiradical effects of resveratrol (RSV), ε-viniferin (VNF), and vitisin B (VB) alone or in combination, and those of a standardized stilbene-enriched vine extract (SSVE). In the DPPH-, FRAP-, and NO-scavenging assays, RSV presented the highest activity with an IC50 of 81.92 ± 9.17, 13.36 ± 0.91, and 200.68 ± 15.40 µM, respectively. All binary combinations resulted in additive interactions in the DPPH- and NO-scavenging assays. In the FRAP assay, a synergic interaction for RSV + VNF, an additive for VNF + VB, and an antagonistic for RSV + VB were observed. The ternary combination of RSV + VNF + VB elicited an additive interaction in the DPPH assay and a synergic interaction in the FRAP- and NO-scavenging assays. There was no significant difference between the antioxidant activity of the SSVE and that of the combination of RSV + VNF. In conclusion, RSV presented the highest effects, followed by VNF and VB. The interactions revealed additive or synergistic effects, depending on the combination of the stilbenes and assay.

Activity of Pterostilbene Metabolites against Liver Steatosis in Cultured Hepatocytes.

Pterostilbene is a dimethyl ether derivative of resveratrol, less metabolized than its analogue, due to the substitution of two hydroxyl groups with methoxyl groups. Nevertheless, the amounts of pterostilbene phase II metabolites found in plasma and tissues are higher than those of the parent compound. The first aim of this study was to assess whether pterostilbene-4'-O-glucuronide (PT-G) and pterostilbene-4'-O-sulfate (PT-S) were able to prevent triglyceride accumulation in AML12 (alpha mouse liver 12) hepatocytes. This being the case, we aimed to analyze the mechanisms involved in their effects. For this purpose, an in vitro model mimicking the hepatocyte situation in fatty liver was developed by incubating mouse AML12 hepatocytes with palmitic acid (PA). For cell treatments, hepatocytes were incubated with 1, 10 or 25 µM of pterostilbene, pterostilbene-4'-O-glucuronide or pterostilbene-4'-O-sulfate for 18 h. Triglycerides and cell viability were assessed by a commercial kit and crystal violet assay, respectively. Protein expression of enzymes and transporters involved in triglyceride metabolism was analyzed by immunoblot. The results showed for the first time the anti-steatotic effect of pterostilbene metabolites and thus, that they contribute to the preventive effect induced by pterostilbene on steatosis in in vivo models. This anti-steatotic effect is mainly due to the inhibition of de novo lipogenesis.

Crown Procyanidin Tetramer: A Procyanidin with an Unusual Cyclic Skeleton with a Potent Protective Effect against Amyloid-ß-Induced Toxicity.

The structure of a new procyanidin tetramer, which we call a crown procyanidin tetramer, with an unprecedented macrocyclic structure has been characterized for the first time. Its comprehensive spectroscopic analysis revealed that it is a symmetric procyanidin tetramer composed of four (-)-epicatechin sub-units linked alternatively via 4ß→8 or 4ß→6 B-type interflavanyl linkages to form the macrocyclic structure. This NMR-characterized carbon skeleton has never been reported before for procyanidins in grape or in wine, neither in the plant kingdom. Surprisingly, the crown procyanidin tetramer appeared to be specifically localized in grape skin, contrasting with the oligomeric and polymeric procyanidins present in seed, skin, and bunch stem. Moreover, this crown procyanidin tetramer showed promising protective effects against amyloid-ß induced toxicity.

In Vitro Glucuronidation and Sulfation of ε-Viniferin, a Resveratrol Dimer, in Humans and Rats.

ε-Viniferin is a resveratrol dimer that possesses antioxidant or anti-inflammatory activities. However little is known about the metabolism of this oligostilbene. This study was thus undertaken as a first approach to identify and characterize the metabolites of ε-viniferin and to describe the kinetic profile of their appearance in humans and rats. The glucuronides and sulfates of ε-viniferin were first obtained by chemical hemi-synthesis and were fully characterized by UPLC-MS and NMR spectroscopy. Then, ε-viniferin was incubated with human or rat S9 liver fractions that led to the formation of four glucuronoconjugates and four sulfoconjugates. In both species, ε-viniferin was subjected to an intense metabolism as 70 to 80% of the molecule was converted to glucuronides and sulfates. In humans, the hepatic clearance of ε-viniferin (Vmax/Km) for glucuronidation and sulfation were 4.98 and 6.35 µL/min/mg protein, respectively, whereas, in rats, the hepatic clearance for glucuronidation was 20.08 vs. 2.59 µL/min/mg protein for sulfation. In humans, three major metabolites were observed: two glucuronides and one sulfate. By contrast, only one major glucuronide was observed in rats. This strong hepatic clearance of ε-viniferin in human and rat could explain its poor bioavailability and could help to characterize its active metabolites.

LC-MS identification and preparative HPLC isolation of Frankenia pulverulenta phenolics with antioxidant and neuroprotective capacities in PC12 cell line.

CONTEXT: Frankenia pulverulenta L. (Frankeniaceae) is a medicinal species with carminative, analgesic and antiviral properties. However, phytochemical investigations, antioxidant and neuroprotective capacities of this plant remain unclear. OBJECTIVE: This work assesses the phenolic composition of F. pulverulenta shoot and root and evaluates their antioxidant and neuroprotective capacities. MATERIALS AND METHODS: Successive fractionation of F. pulverulenta shoot and root using 6 solvents were used. Antioxidant capacity of these fractions was assessed through four in vitro tests (DPPH, ABTS, Fe-chelating activity and ORAC). Phenolic identification, purification as well as neuroprotective activity of ethyl acetate (EtOAc) fraction and purified molecules were assessed. RESULTS: Among the tested fractions, EtOAc shoot and root fractions possessed considerable phenolic contents (383 and 374 mg GAE/g E, respectively) because of their important ORAC (821 and 1054 mg of TE/g E), DPPH (586 and 750 mg of TE/g) and ABTS (1453 and 1319 mg of TE/g) results. Moreover, gallic acid, quercetin, quercetin galloyl glucoside, trigalloyl hexoside, procyanidin dimers and sulfated flavonoids were identified by LC-DAD-ESI-MS for the first time in this species. The relevant cytoprotective capacity (at 300 µg/mL) against ß-amyloid peptide induced toxicity in PC12 cells of EtOAc fractions were corroborated with the chemical composition. In addition, purified molecules were tested for their ORAC and neuroprotective activity. Quercetin showed the best ORAC value (33.55 mmol TE/g polyphenols); nevertheless, procyanidin dimer exhibited an exceptionally efficient neuroprotective activity (100% of viability at 50 µg/mL). DISCUSSION AND CONCLUSIONS: These findings suggest that this halophyte is a promising source of antioxidant and neuroprotective molecules for pharmaceutical purposes.

Two New Stilbenoids from the Aerial Parts of Arundina graminifolia (Orchidaceae).

Two new phenanthrene derivatives, a phenanthrenequinone named arundiquinone (1) and a 9,10-dihydrophenanthrene named arundigramin (2) together with a known lignin dimer (3) and seven known stilbenoids (4-10) were isolated from the aerial parts of the Asian orchid Arundina graminifolia. The structures of the isolated compounds were elucidated by spectroscopic methods, including extensive 1D, 2D NMR (heteronuclear single quantum coherence (HSQC), heteronuclear multiple-bond correlation spectroscopy (HMBC), and HR-ESI-MS techniques, as well as comparison with respective literature reports. The cytoprotective activity of the isolated compounds were evaluated for their ability to reduce beta amyloid induced toxicity on undifferentiated PC12 cells. Compound 8 showed moderate cytoprotective activity at 0.5 µmol/L (71% of cell viability) while the other compounds showed no significant activity at the highest concentration tested.

Viniphenol A, a complex resveratrol hexamer from Vitis vinifera stalks: structural elucidation and protective effects against amyloid-ß-induced toxicity in PC12 cells.

Stilbenes have received much attention during the last two decades following the discovery of resveratrol in wine. Since then, there have been a growing number of papers reporting various biological activities of naturally occurring stilbenes. The aim of this study was to determine new minor stilbenes from Vitis vinifera stalks. Purification of these compounds was achieved by means of centrifugal partition chromatography, a versatile separation technique that does not require a solid stationary phase. Viniphenol A (1), a new resveratrol hexamer, was isolated along with five oligostilbenoids identified in V. vinifera for the first time, ampelopsin C, davidiol A, leachianol F, leachianol G, and E-maackin, a dimer with an unusual dioxane moiety, and 14 known hydroxystilbenes. The structure and stereochemistry of viniphenol A were determined on the basis of spectroscopic data analysis and molecular modeling under NMR constraints. Viniphenol A showed protective effects against amyloid-ß-induced toxicity in PC12 cell cultures.

Characterization of Crocetin Isomers in Serum Samples via UHPLC-DAD-MS/MS and NMR after Saffron Extract (Safr'Inside™) Consumption.

The therapeutic effects of saffron have been reported and described in relation to its major derivatives. Among them, in terms of saffron's properties, crocin and crocetin absorption and bioavailability have been the most studied. Nevertheless, the metabolism of these major compounds of saffron has not yet been entirely elucidated. Current data indicate that the phase 2 metabolism of crocetins go through conjugation reactions. Crocetins could also be present in isomeric forms such as other carotenoids. Nonetheless, there are still shadow areas in regard to the measurements of the different circulating forms of crocetins after oral saffron extract administration (Safr'Inside™). In using various approaches, we propose the identification of a new cis isomeric form of crocetin, the 6-cis-crocetin. This compound was found in human serum samples after an oral administration of saffron extract. The 6-cis-crocetin represents 19% of the total crocetin measured after 45 min of consumption. These data mark, for the first time, the presence of a cis isomeric form of crocetin in human serum samples. Moreover, this study led to the development of an analytical method that is able to identify and quantify both isomeric forms (trans and cis).

Reduced Production of Pro-Inflammatory and Pro-Catabolic Factors by Human Serum Metabolites Derived from a Patented Saffron Extract Intake.

Safe and anti-inflammatory plant-based natural products present an increasing focus in the treatment of chronic inflammatory diseases such as osteoarthritis or inflammatory bowel diseases. Among them, saffron, a spice derived from the stigma of Crocus sativus, could have anti-inflammatory properties and would be therefore a promising therapeutic agent for the treatment of such conditions. However, the anti-inflammatory molecular mechanisms of saffron in humans are still understudied and unclear. In this study, combining human serum metabolites and cell cultures, we evaluated the effect of circulating metabolites from the consumption of a patented saffron extract (Safr'InsideTM) on the chondrocytes and colon epithelial cell responses to inflammatory stress. Parametric or non-parametric Analysis of Variance with post hoc tests was performed. We demonstrated that human serum containing metabolites from saffron intake attenuated IL-1ß-stimulated production of PGE2 and MMP-13 in chondrocyte cells and limited the increase in ICAM-1, MCP-1, iNOS, and MMP-3 in human epithelial cells following combined IL-1ß and TNF-α inflammatory stimulation. Altogether, these data provide new findings into the mechanisms underlying the beneficial effects of saffron on chondrocytes and enterocyte cells at the cellular level and in the context of chronic inflammatory disorders.

Inhibitory activity of plant stilbenoids against nitric oxide production by lipopolysaccharide-activated microglia.

Microglia-driven inflammatory processes are thought to play an important role in ageing and several neurological disorders. Since consumption of a diet rich in polyphenols has been associated with anti-inflammatory and neuroprotective effects, we studied the effects of twenty-five stilbenoids isolated from Milicia excelsa, Morus alba, Gnetum africanum, and Vitis vinifera. These compounds were tested at 5 and 10 µM on BV-2 microglial cells stimulated with bacterial lipopolysaccharide. Ten stilbenoids reduced lipopolysaccharide-induced nitric oxide production at 5 and/or 10 µM. Two tetramers, E-vitisin A and E-vitisin B, were the most effective molecules. Moreover, they attenuated the expression of the inducible NO synthase protein and gene.

Prediction of the penetration depth of multi-lamellar liposomes in artificial skin. Application to the vectorization of corticosteroid in human skin.

Our previous work showed that the size, elasticity and charge of multi-lamellar liposomes (MLLs) could not be considered separately to predict the fate of MLLs in the skin [1]. Based on this study, we developed several MLLs formulations containing a corticosteroid, betamethasone 17-valerate (B17) to transport the drug into the stratum corneum, living epidermis, dermis or through the skin. MLLs encapsulation efficiency was found to exceed 74 ± 3 % in all cases. In addition, we showed that MLLs protected the corticosteroid from thermal degradation. Comparing the penetration depth of all MLLs within artificial skin measured by Raman imaging, we established an equation for its determination, given the MLLs elasticity and size. This equation was verified experimentally on human explants: quantification of B17 in each skin layer, as well as its transdermal passage by ultra-high performance liquid chromatography, confirmed that B17 was predominantly and significantly transported in the desired layer. Eventually, we showed the benefits in using B17-loaded MLLs instead of a B17-containing pharmaceutical cream in terms of B17 penetration and thermal degradation.

Beneficial Effects of ε-Viniferin on Obesity and Related Health Alterations.

Viniferin is a phenolic compound belonging to the group of stilbenoids. In particular, ε-viniferin is a dimer of resveratrol, found in many plant genders, among which grapes (Vitis vinifera) are a primary source. Due to the fact that ε-viniferin is mainly present in the woody parts of plants, their use as a source of this bioactive compound is a very interesting issue in a circular economy. Both, in vitro studies carried out in pre-adipocytes and mature adipocytes and in vivo studies addressed in mice show that ε-viniferin is able to reduce fat accumulation. Moreover, it prevents the development of some obesity co-morbidities, such as type 2 diabetes, dyslipidemias, hypertension and fatty liver. ε-viniferin can be absorbed orally, but it shows a very low bioavailability. In this scenario, further research on animal models is needed to confirm the effects reported in a great number of studies; to determine which metabolites are involved, including the main one responsible for the biological effects observed and the mechanisms that justify these effects. In a further phase, human studies should be addressed in order to use ε-viniferin as a new tool for obesity management, as a nutraceutical or to be included in functional foods.

Circulating Human Metabolites Resulting from TOTUM-070 Absorption (a Plant-Based, Polyphenol-Rich Ingredient) Improve Lipid Metabolism in Human Hepatocytes: Lessons from an Original Ex Vivo Clinical Trial.

TOTUM-070 is a patented polyphenol-rich blend of five different plant extracts showing separately a latent effect on lipid metabolism and potential synergistic properties. In this study, we investigated the health benefit of such a formula. Using a preclinical model of high fat diet, TOTUM-070 (3 g/kg of body weight) limited the HFD-induced hyperlipemia with a reduction in triglyceride (-32% after 6 weeks; -20.3% after 12 weeks) and non-HDL cholesterol levels (-21% after 6 weeks; -38.4% after 12 weeks). To further investigate such a benefit and its underlying mechanisms in humans, we designed an ex vivo clinical approach to collect the circulating bioactives resulting from TOTUM-070 ingestion and to determine their biological activities on human hepatocytes. Human serum was obtained from healthy subjects before and after intake of TOTUM-070 (4995 mg). The presence of circulating metabolites was assessed by UPLC-MS/MS. Serum containing metabolites was further incubated with hepatocytes cultured in a lipotoxic environment (palmitate, 250 µM). RNA sequencing analyses show that lipid metabolism was one of the most impacted processes. Using histologic, proteomic, and enzymatic assays, the effects of human TOTUM-070 bioactives on hepatocyte metabolism were characterized by (1) the inhibition of lipid storage, including both (2) triglycerides (-41%, p < 0.001) and (3) cholesterol (-50%, p < 0.001) intracellular content, (4) a reduced de novo cholesterol synthesis (HMG-CoA reductase activity -44%, p < 0.001), and (5) a lowered fatty acid synthase protein level (p < 0.001). Altogether, these data support the beneficial impact of TOTUM-070 on lipid metabolism and provide new biochemical insights in human mechanisms occurring in liver cells.

In the shadow of resveratrol: biological activities of epsilon-viniferin.

Stilbenes are secondary metabolites belonging to the polyphenol family. Those compounds are derived from the glycosylation, prenylation, methoxylation, hydroxylation, or also oligomerization of the well-known trans-resveratrol. One of them, trans-epsilon-viniferin (ε-viniferin), is a trans-resveratrol dimer that arouses the interest of researchers in the field of human health. The biosynthesis of this molecule in various plant species, particularly high in the Vitaceae family, explains its presence in some red wines, which represent the main source of ε-viniferin in the human diet. Although bioavailability studies have shown poor absorption and high metabolism of this stilbene, multiple studies demonstrated its biological properties. The ε-viniferin exhibits strong activities against inflammatory and oxidative stress. Moreover, various studies have reported great activity of this compound not only in a wide range of disorders and diseases, such as cancer, obesity, and its associated disorders, but also in vascular diseases and neurodegeneration, for which the pathophysiology is closely related to the state of oxidation and inflammation. This review provides a state of art of the main activities of ε-viniferin demonstrated in vitro and in vivo, highlighting that this resveratrol dimer could be a promising candidate for future functional foods or supplement foods used for the management of many chronic diseases of concern in terms of public health.

Oxyresveratrol and Gnetol Glucuronide Metabolites: Chemical Production, Structural Identification, Metabolism by Human and Rat Liver Fractions, and In Vitro Anti-inflammatory Properties.

Stilbene metabolites are attracting great interest because many of them exhibit similar or even stronger biological effects than their parent compounds. Furthermore, the metabolized forms are predominant in biological fluids; therefore, their study is highly relevant. After hemisynthesis production, isolation, and structural elucidation, three glucuronide metabolites for oxyresveratrol (ORV) were formed: trans-ORV-4'-O-glucuronide, trans-ORV-3-O-glucuronide, and trans-ORV-2'-O-glucuronide. In addition, two glucuronide metabolites were obtained for gnetol (GN): trans-GN-2'-O-glucuronide and trans-GN-3-O-glucuronide. When the metabolism of ORV and GN is studied in vitro by human and rat hepatic enzymes, four of the five hemisynthesized compounds were identified and quantified. Human enzymes glucuronidated preferably at the C-2' position, whereas rat enzymes do so at the C-3 position. In view of these kinetic findings, rat enzymes have a stronger metabolic capacity than human enzymes. Finally, ORV, GN, and their glucuronide metabolites (mainly at the C-3 position) decreased nitric oxide, reactive oxygen species, interleukin 1ß, and tumor necrosis factor α production in lipopolysaccharide-stimulated macrophages.

Protective effect of ε-viniferin on ß-amyloid peptide aggregation investigated by electrospray ionization mass spectrometry.

Abnormal ß-amyloid peptide accumulation and aggregation is considered to be responsible for the formation and cerebral deposition of senile plaques in the brains of patients with Alzheimer's disease (AD). Inhibition of the formation of ß-amyloid (Aß) fibrils would be an attractive therapeutic target for the treatment of AD. Resveratrol and its derivatives exhibit a broad range of pharmacological properties such as protection against cardiovascular diseases and cancers, as well as promoting antiaging effects. We reported previously that ε-viniferin glucoside (VG), a resveratrol-derived dimer, strongly inhibits Aß (25-35) fibril formation in vitro. In this study, we investigated the effects of VG on the aggregation of the full-length peptides (Aß (1-40) and Aß (1-42)) and on the ß-amyloid-induced toxicity in PC12 cells. VG inhibited Aß cytotoxicity and the non-covalent complex between VG and Aß was observed by electrospray ionization mass spectrometry.

Encapsulation of ε-Viniferin into Multi-Lamellar Liposomes: Development of a Rapid, Easy and Cost-Efficient Separation Method to Determine the Encapsulation Efficiency.

Onion-type multi-lamellar liposomes (MLLs), composed of a mixture of phosphatidylcholine and Tween 80, were analyzed for their ability to encapsulate ε-Viniferin (εVin), a resveratrol dimer. Their encapsulation efficiency (EE) was measured by UV-VIS spectroscopy using three different separation methods-ultracentrifugation, size exclusion chromatography, and a more original and advantageous one, based on adsorption filtration. The adsorption filtration method consists indeed of using syringe filters to retain the molecule of interest, and not the liposomes as usually performed. The process is rapid (less than 10 min), easy to handle, and inexpensive in terms of sample amount (around 2 mg of liposomes) and equipment (one syringe filter is required). Whatever the separation method, a similar EE value was determined, validating the proposed method. A total of 80% ± 4% of εVin was found to be encapsulated leading to a 6.1% payload, roughly twice those reported for resveratrol-loaded liposomes. Finally, the release kinetics of εVin from MLLs was followed for a 77 day period, demonstrating a slow release of the polyphenol.

In Vivo Genotoxicity Evaluation of a Stilbene Extract Prior to Its Use as a Natural Additive: A Combination of the Micronucleus Test and the Comet Assay.

Genotoxic data of substances that could be used as food additives are required by the European Food Safety Authority. In this sense, the use of an extract from grapevine shoots containing a stilbene richness of 99% (ST-99), due to its antioxidant and antibacterial activities, has been proposed as an alternative to sulfur dioxide in wine. The aim of this work was to study, for the first time, the in vivo genotoxic effects produced in rats orally exposed to 90, 180, or 360 mg ST-99/kg body weight at 0, 24, and 45 h. The combination of micronucleus assay in bone marrow (OECD 474) and standard (OECD 489) and enzyme-modified comet assay was used to determine the genotoxicity on cells isolated from stomach, liver, and blood of exposed animals. The ST-99 revealed no in vivo genotoxicity. These results were corroborated by analytical studies that confirm the presence of stilbenes and their metabolites in plasma and tissues. Moreover, to complete these findings, a histopathological study was performed under light microscopy in liver and stomach showing only slight modifications in both organs at the highest concentration used. The present work confirms that this extract is not genotoxic presenting a good profile for its potential application as a preservative in the wine industry.