Hervé Rogez

Optimisation of the solvent extraction of phenolic antioxidants from fruits of Euterpe oleracea using Response Surface Methodology.

Fruits of Euterpe oleracea (FEO) are currently known as elements that present a very high antioxidant activity (AAO), as measured by the Oxygen Radical Absorbance Capacity. They are particularly rich in total phenolics (TP) and total anthocyanins (TA). Response Surface Methodology was used to optimise the solvent extraction of phenolic antioxidants from FEO, using a second-order polynomial equation to describe the experimental data for TP, TA, and AAO. In order to determine the best solid-to-liquid ratio and time of extraction, some preliminary studies were conducted. A rotatable central composite design with three variables (ethanol proportion, hydrochloric acid concentration and temperature) was then used. The results showed a good fit to the proposed model (R(2)>0.89). TP and TA, as well as TA and AAO, showed significant correlations (P<0.05). The optimised conditions that maximized the yields of phenolic compounds (TP and TA) and AAO from FEO were: ethanol proportion between 70% and 80%, hydrochloric acid concentration between 0.065 and 0.074 mol/L and a temperature of 58 degrees C.

Piceatannol, a potent bioactive stilbene, as major phenolic component in Rhodomyrtus tomentosa

The sim fruit (Rhodomyrtus tomentosa) has long been used in folk medicine to treat diarrhoea, dysentery, and to boost the immune system. The purpose of this work was to determine its phenolic profile and to evaluate the changes of content during maturation, as well as the variations induced by environmental conditions. Using HPLC-ESI-HR-MS, 19 phenolic compounds (PCs) were tentatively characterised and included stilbenes and ellagitannins as major components, followed by anthocyanins, flavonols, and gallic acid. PCs were then further quantified by HPLC-DAD. Piceatannol, a promising health-promoting stilbene component, was the major PC in the fruit with a concentration of 2.3mg/g dry weight at full maturity stage. This concentration is 1000-2000 times higher than that of red grapes, a major source of stilbene in the human diet. During maturation, the contents in piceatannol and other stilbenes, ellagitannins, and flavonols decreased while the anthocyanin content increased. Shade-grown sim fruits showed significantly higher piceatannol levels than sun-exposed fruits. Taken together, these findings highlight the potential of sim, an under-utilised plant species from South-East Asia, as a source of health-promoting fruits.

Development of a standardised human in vitro digestion protocol based on macronutrient digestion using response surface methodology.

Bioaccessibility studies should be taken into account when evaluating the physiological effects of ingested compounds at the intestine level. Several in vitro digestion protocols have been described, with a wide range of experimental conditions but no optimised protocol exists. In order to fill in this gap, we evaluated the influence of three continuous factors (pH, incubation time, and enzyme concentrations), in the range of values found in literature, on the digestion of standard macronutrients (starch, albumin, triolein) alone or in mixture. Three central composite designs, using response surface methodology, were employed to model the three abiotic steps of pre-colonic digestion. A validated in vitro digestion was eventually set up for the salivary step (pH 6.9, 5 min, 3.9 units α-amylase/ml), the gastric step (pH 2, 90 min, 71.2 units pepsin/ml), and the abiotic duodenal step (pH 7, 150 min, 9.2mg pancreatin and 55.2mg bile extract/ml).

Phenolic profiling in the pulp and peel of nine plantain cultivars (Musa sp)

The present study investigated the phenolic profiles of the pulp and peel of nine plantain cultivars and compared them to those of two dessert bananas of commercial interest (Grand Nain and Gros Michel), alongside a newly created hybrid, resistant to black sigatoka disease (F568). Identification and quantification of phenolic compounds were performed by means of HPLC-ESI-HR-MS and HPLC-DAD. Hydroxycinnamic acids, particularly ferulic acid-hexoside with 4.4-85.1 μg/g of dry weight, dominated in the plantain pulp and showed a large diversity among cultivars. Flavonol glycosides were predominant in plantain peels, rutin (242.2-618.7 μg/g of dry weight) being the most abundant. A principal component analysis on the whole data revealed that the phenolic profiles of the hybrid, the dessert bananas and the pure plantains differed from each other. Plantain pulps and peels appeared as good sources of phenolics, which could be involved in the health benefits associated with their current applications.

A rapid validated UHPLC–PDA method for anthocyanins quantification from Euterpe oleracea fruits

The aim of this work is to develop the first validated UHPLC-PDA method for major anthocyanins quantification in Euterpe oleracea fruits after fast extraction procedures and samples preparation. The separation was performed on HSS C18 column (1.8 μm) using a gradient elution with acetonitrile and 5% formic acid in a total run time of only 17 min. Total error and accuracy profiles were used as criteria for the validation process. Calibration in the matrix was found to be more accurate than calibration without matrix. Trueness (<6.76% relative bias), repeatability (<4.6% RSD), intermediate precision (<5.3% RSD), selectivity, response function and linearity for major anthocyanins, cyanidin-3-glucoside and cyanidin-3-rutinoside, were evaluated. The concentration range validated was 1-48 μg/mL for both compounds. In addition two cyanidin-di-O-glycosides were detected for the fist time in this fruit. We also showed that a first extraction of the fruits with ethyl acetate removes the lipophilic compounds and allows an easier extraction by methanol and quantification of anthocyanins in this extract.

Linseed oil stabilisation with pure natural phenolic compounds

Linseed has been used for a very long time in human and animal nutrition. Currently, there is an increasing interest in linseed oil because of its particularly high content in α-linolenic acid (ALA), an omega-3 fatty acid (FA). Unfortunately, ALA turns also the oil extremely sensitive to oxidation. This study aimed at assessing four pure representative phenolic compounds, myricetin (flavonol), (+)-catechin (flavanol), genistein (isoflavone), and caffeic acid (hydroxycinnamic acid) at a concentration of 555μmol/kg as antioxidants in refined linseed oil (RLO). Their protective effect was assessed by monitoring the hydroperoxide formation, the FA profile and the residual antioxidant concentration in RLO, along its storage at 60°C according to the Schaal oven test procedure. Caffeic acid, (+)-catechin and myricetin were found to be more efficient than butylated hydroxyanisole (BHA), a synthetic antioxidant. Interestingly, myricetin strongly reduced ALA oxidation. These results confirm that the chemical structure of the phenolic compounds plays a major role in their antioxidant properties.

Nutritional composition and antioxidant properties of the sim fruit (Rhodomyrtus tomentosa)

In this study, detailed chemical properties of sim (Rhodomyrtus tomentosa (Ait.) Hassk.) fruit including nutritional composition, phenolic content and antioxidant capacity were determined for the first time. A 150g serving of sim fruit contained high levels of dietary fibre (69.94-87.43% of Recommended Daily Intake (RDI)), α-tocopherol (38.90-51.87% RDI), manganese (>100% RDI), and copper (44.44% RDI) but low levels of protein (2.63% RDI), lipid (1.59-3.5% RDI), and sugars (5.65% RDI). The predominant fatty acid in the sim fruit sample was linoleic acid (75.36% of total fatty acids). Interestingly, total phenolics (49.21±0.35mg gallic acid equivalent (GAE)/g dry weight (DW)) were particularly high and resulted in a high antioxidant capacity (431.17±14.56μmol Trolox equivalent (TE)/g DW). These results, together with our recent discovery of high amount of piceatannol, a stilbene with potent biological activities, highlight the potential of sim, an under-utilised plant species from South-East Asia, as a new source of health-promoting compounds including dietary fibres, essential fatty acids, and phenolic compounds.

Identification and antioxidant activity of several flavonoids of Inga edulis leaves

Um extrato metanol-agua das folhas de Inga edulis foi fracionado para identificar os compostos polifenolicos. Os compostos identificados foram o acido galico, a catequina, a epicatequina, a miricetina-3-ramnopiranosideo, a quercetina-3-glucopiranosideo e a quercetina-3-ramnopiranosideo. A capacidade antioxidante do extrato e dos polifenois puros foi medida pelo teste ORAC e comparada com o teor em fenolicos totais (TP). O extrato bruto seco apresentou valores de ORAC (11.16 mmol TE per g) e TP (496.5 mg GAE per g) muito altos. Os compostos identificados foram responsaveis, respectivamente, por 9.53 % e 12.10 % dos valores ORAC e de TP do extrato de folhas de Inga edulis.

Enriquecimento de compostos fenólicos de folhas de Inga edulis por extração em fase sólida: quantificação de seus compostos majoritários e avaliação da capacidade antioxidante

A phenolic fraction was obtained from of the acetone-water-acetic acid extract of Inga edulis leaves, by liquid-liquid partition and SPE-C18 cartridges. This method provided an increase of 108, 66, 51, 50 and 36% of flavonols, proanthocyanidins, total polyphenols, gallotannins and flavanols, respectively. The major phenolics in purified fraction were procyanidin B2, catechin and myricetin-3-O-α-L-rhamnopyranoside, which achieved increases of 111, 47 and 45%, respectively, after SPE. Acid hydrolysis confirmed the presence of procyanidins, prodelphinidins and glycosylated flavonoids.

Immunomodulatory action of Copaifera spp oleoresins on cytokine production by human monocytes

Copaifera spp oleoresins have been used in folk medicine for centuries; nevertheless, its immunomodulatory action has not been investigated. Thus, the goal of this study was to characterize different oleoresins and to verify their action on human monocytes regarding pro- and anti-inflammatory cytokine production (TNF-α and IL-10, respectively). The chemical composition of Brazilian Copaifera reticulata, Copaifera duckey and Copaifera multijuga oleoresins was analyzed by HPLC-MS. Cell viability was assessed by MTT method after incubation of cells with Copaifera spp. Noncytotoxic concentrations of oleoresins were incubated with human monocytes from healthy donors, and cytokine production was determined by ELISA. HPLC-MS analysis for terpenes allowed the identification of six diterpene acids and one sesquiterpene acid. Oleoresins exerted no cytotoxic effects on human monocytes. All oleoresins had a similar profile: LPS-induced TNF-α production was maintained by oleoresins, while a significant inhibitory action on IL-10 production was seen. Copaifera oleoresins seemed to exert an activator profile on human monocytes without affecting cell viability. Such effect may be due to the presence of either diterpene or sesquiterpene acids; however, further studies are necessary to determine the involvement of such compounds in Copaifera immunomodulatory effects.