Bio-fortification and isotopic labelling of Se metabolites in onions and carrots following foliar application of Se and Se

The aims were to bio-fortify onions by foliar application of selenium (Se) and to intrinsically label bioactive Se-metabolites in onion and carrot by enriched, stable ⁷⁷Se for use in human physiological studies. Onion bulbs and leaves were enriched in Se by repeated foliar spraying of 10 or 100 μg Se ml⁻¹ solutions of sodium selenite (Se(IV)) or sodium selenate (Se(VI)). ICP-MS analysis of onion leaves and bulbs showed that the Se concentration was enhanced by up to a factor of approximately 50 and 200 in bulbs and leaves, respectively. HPLC–ICP-MS analysis of proteolytic plant extracts showed that foliar application of Se(IV) gave rise to bio-synthesis of a higher fraction of the desired organic Se species and was better tolerated by the plants than Se(VI). Based on these findings onions and carrots were bio-fortified by foliar application of a solution of ⁷⁷Se(IV) that was enriched to 99.7% as ⁷⁷Se. The ⁷⁷Se- labelled metabolites in onions were predominantly γ-glutamyl-⁷⁷Se-selenomethyl-selenocysteine (γ-glu-Me⁷⁷SeCys), ⁷⁷Se-methylselenocysteine (Me⁷⁷SeCys) and ⁷⁷Se-selenomethionine (⁷⁷SeMet). Furthermore, we report here for the first time the finding in carrots of the bioactive Me⁷⁷SeCys, the identity of which was verified by HPLC–ESI-MS/MS.     

Selenium species in buckwheat cultivated with foliar addition of Se(VI) and various levels of UV-B radiation

Common (Fagopyrum esculentum Moench) and tartary (Fagopyrum tataricum Gaertn.) buckwheat was treated by spraying the leaves with a water solution containing 15 mg Se per litre in the form of sodium selenate in the flowering period. The selenium content in all parts of plant was found to be less than 200 ng g⁻¹ in non-treated and in the range 2700–4650 ng g⁻¹ in selenium treated buckwheat. Exposure to UV-B radiation lead to higher Se accumulation in flowers of both Se enriched cultivars. For speciation analysis enzymatic hydrolysis was carried out, separation and detection of selenium species was performed by high performance liquid chromatography–ultraviolet treatment–hydride generation atomic fluorescence spectrometry (HPLC–UV–HG-AFS). In flowers and leaves, on average 11% of the Se content was soluble and in the form of Se(VI), representing between 0.6% (flowers) and 3% (leaves) of the Se content. The remaining soluble non-amino acid organic Se was not detected by HPLC–UV–HG-AFS. In seeds 93% of the selenium content was found in the extracts and the main selenium species was SeMet with 93 ± 5% relative to the selenium content.     

Supercritical fluid extraction of daidzein and genistein isoflavones from soybean hypocotyl after hydrolysis with endogenous β-glucosidases

An optimal condition of supercritical fluid extraction (SFE) for isoflavone aglycones (daidzein and genistein) in soybean hypocotyls previously subjected to thermohydration at pH 5.0 and a temperature of 50 °C for 6, 12 and 18 h was developed. Different temperatures, pressures and cosolvents (methanol, ethanol, and acetonitrile) was tested and compared with solid–liquid extraction using aqueous methanol solution (80% v v⁻¹) conducted in parallel for comparison. The extraction conditions were 50–70 °C, 176–380 bar, adding 0, 5, 10 mol% of cosolvents 80% in water as a modifier. The results from SC–CO₂ showed that the cosolvent and pressure have significant effects in the extraction efficiency. It was found that the extraction conditions promoting the highest extraction of daidzein and genistein were at the temperature of 60 °C, pressure of 380 bar and both static and dynamic extraction of 15 min with the addition of 10% acetonitrile (80% v v⁻¹). The maximum amounts of daidzein and genistein extracted by each method were solid–liquid extraction (70.07 mg 100 g⁻¹) and carbon dioxide–acetonitrile (17.97 mg 100 g⁻¹). The yield of daidzein and genistein achieved by a 30 min SC–CO₂ extraction on soybean hypocotyls after 12 h soaking time was markedly improved by the addition of a modifier (acetonitrile) to the CO₂ fluid. HPLC analysis of the obtained extracts revealed that extraction of isoflavone aglycones by SC–CO₂ was 4.78 and 13.19 mg 100 g⁻¹ for daidzein and genistein, respectively. The contents of daidzein and genistein obtained in the solid–liquid extraction were superior to 86% and 63%, respectively, compared to supercritical extraction.     

Determination of selenium in selected food samples from Argentina and estimation of their contribution to the Se dietary intake

An optimised FI-HGAAS method was applied to determine the total selenium concentrations in selected high consumption food (fish, beef, chicken, milk, rice, wheat flour, egg) and to estimate their contribution to the Argentinean dietary intake, whose information is scarce nowadays. Through several optimisation steps a suitable method was achieved showing satisfactory figures of merit for all matrices. Average recovery was 96%, RSD < 5%, LODs ranged 2.0–7.0 μg kg⁻¹ and the accuracy was assessed using DOLT-3 NRC certified reference material. Meat and eggs showed the highest values (in μg kg⁻¹, beef: 42–153; chicken: 62–205; fish: 94–314; canned tuna: 272–282; eggs: 134–217), minor values were found for wheat flour (22–42), rice: (<22), pasta (47–64) and milk (<7–9). An estimated intake of 32 and 24 μg day⁻¹ for adult men and women, respectively, suggested a deficient Se intake, leading to further comprehensive surveys of Se occurrence in Argentina.     

Effect of foliar application of selenium on its uptake and speciation in carrot

Carrot (Daucus carota) shoots were enriched by selenium using foliar application. Solutions of sodium selenite or sodium selenate at 10 and 100 μg Se ml⁻¹, were sprayed on the carrot leaves and the selenium content and uptake rate of selenium were estimated by ICP–MS analysis. Anion and cation exchange HPLC were tailored to and applied for the separation of selenium species in proteolytic extracts of the biological tissues using detection by ICP–MS or ESI–MS/MS. Foliar application of solutions of selenite or selenate at 100 μg Se ml⁻¹ resulted in a selenium concentration of up to 2 μg Se g⁻¹ (dry mass) in the carrot root whereas the selenium concentration in the controls was below the limit of detection at 0.045 μg Se g⁻¹ (dry mass). Selenate-enriched carrot leaves accumulated as much as 80 μg Se g⁻¹ (dry mass), while the selenite-enriched leaves contained approximately 50 μg Se g⁻¹ (dry mass). The speciation analyses showed that inorganic selenium was present in both roots and leaves. The predominant metabolised organic forms of selenium in the roots were selenomethionine and γ-glutamyl-selenomethyl-selenocysteine, regardless of which of the inorganic species were used for foliar application. Only selenomethionine was detected in the carrot leaves. The identity of selenomethionine contained in carrot roots and leaves was successfully confirmed by HPLC–ESI–MS/MS.     

Response surface modelling of lead pre-concentration from food samples by miniaturised homogenous liquid–liquid solvent extraction: Box–Behnken design

In this study, a new method called miniaturised homogenous liquid–liquid extraction, followed by graphite furnace atomic absorption spectrometry, was developed for the extraction and determination of lead from food samples. The procedure was based on the fast extraction of lead from an acetic acid sample solution into 0.5 mL chloroform, as an extraction solvent. After adding water into the mixture, the extracting solvent phase immediately formed a distinct water-immiscible phase below the vial, which could easily be separated, evaporated and re-dissolved in 1.0 mL nitric acid 0.1 mol L⁻¹ for further analysis. The effects of various experimental parameters in extraction step were studied using two optimisation methods, one variable at a time and Box–Behnken design. The results showed that the amount of salt and extraction time did not have effect on the extraction efficiency. Therefore, a three-level Box–Behnken experimental design with three factors, which combined the response surface modelling, was used to optimise lead extraction. Three independent variables, including pH of solution (ranging from 6.5 to 10.5), concentration of dithizone as chelating agent (ranging from 0.05 to 0.5 μg L⁻¹) and extracting solvent volume (ranging from 300 to 900 μL) were respectively coded as pH, D and V at three different levels (−1, 0 and 1). In this study, the optimum condition was determined at pH 8.4, a volume of chloroform at 0.45 mL, and concentration of dithizone at 0.5 μg L⁻¹. Under the optimum condition, the limit of detection (LOD) was 0.05 μg L⁻¹. Furthermore, the relative standard deviation of the ten replicate was <5.0%. The developed procedure was applied to the extraction and determination of lead in the food samples.     

Se improves indole glucosinolate hydrolysis products content, Se-methylselenocysteine content, antioxidant capacity and potential anti-inflammatory properties of sauerkraut

Selenium (Se) has a well-known role in prevention of chronic diseases associated with oxidative stress and inflammation. The objective was to produce Se-enriched sauerkraut and to study the effect of selenite addition on indole glucosinolate hydrolysis products, vitamin C, Se biotransformation and microbial quality. The antioxidant activity of sauerkraut and its ability to inhibit NO production in LPS-induced macrophages were also examined. White cabbage was naturally fermented with 0.3 mg of Na₂SeO₃/kg fresh cabbage (NFSe) or without Se addition (NF). Total Se content reached up to 1.29 μg/g d.m. (0.11 μg/g f.m.). Selenomethylselenocysteine was the major Se form found in NFSe cabbage. Se addition caused a slight reduction of ascorbigen (6%) and vitamin C (5%) contents in sauerkraut (P ? 0.05); however, lactic acid bacteria increased (3%), and the formations of indole-3-carbinol and indole-3-acetonitrile were markedly enhanced (74% and 13%, respectively). NFSe cabbage extracts showed higher (P ? 0.05) antioxidant activity (163 μmol trolox/g d.m.) and inhibition of NO production in LPS-induced macrophages (IC₅₀ = 44.0 μg/ml) than did NF cabbage extracts. Consequently, Se-enriched sauerkraut can be considered as a health-promoting food.     

Dispersive liquid–liquid microextraction for the determination of organochlorine pesticides residues in honey by gas chromatography-electron capture and ion trap mass spectrometric detection

A simple dispersive liquid–liquid microextraction (DLLME) protocol for the determination of 15 organochlorine pesticides residues in honey is proposed. The selected pesticides were separated using gas chromatography and detected by electron capture (ECD) or ion trap mass spectrometry (GC-IT/MS). Several parameters affecting the extraction efficiency namely type and volume of organic extraction solvent, type and volume of disperser solvent, sample pH, ionic strength, extraction time and centrifugation speed were systematically investigated. The final DLLME protocol involved the addition of 750 μL acetonitrile (disperser) and 50 μL chloroform (extraction solvent) into a 5 mL aqueous honey solution followed by centrifugation. The sedimented organic phase (chloroform) were analysed directly by GC-IT/MS or evaporated and reconstituted in acetonitrile prior to the GC-ECD analysis. The analytical performance of the GC-ECD and GC-IT/MS methods was compared and discussed. Under the selected experimental conditions, the enrichment factors varied between of 36 and 114. The limits of detection (LOD) were in the range of 0.02–0.15 μg L⁻¹ (0.4–3 ng g⁻¹) for GC-ECD and 0.01–0.2 μg L⁻¹ (0.2–4 ng g⁻¹) for GC-IT/MS which is adequate to verify compliance of products to legal tolerances. The proposed method was applied to the analysis of the selected organochlorine pesticides residues in various honey samples obtained from Greek region. Mean recoveries were ranged from 75% to 119% while the precision was better than 20% in both methodologies.     

Electrocatalysis and sensitive determination of Sudan I at the single-walled carbon nanotubes and iron(III)-porphyrin modified glassy carbon electrodes

Iron-porphyrin (5,10,15,20-tetraphenyl-21H, 23H-porphine iron(III) chloride) was used in combination with single-wall carbon nanotubes (SWNTs) to modify a glassy carbon electrode (GCE). The electrochemical behavior of Sudan I on the iron-porphyrin-SWNT-DMF (N,N-dimethylformamide) modified GCE was studied by the cyclic voltammetry and square wave voltammetry. In pH 7.0 Tris–HCl buffers, Sudan I has a sensitive catalytic reduction peak at −0.08 V on the iron-porphyrin-SWNT-DMF modified GCE. Using square wave voltammetry, the linear relationship of Sudan I is 5.03 × 10⁻⁸ mol L⁻¹–2.01 × 10⁻⁶ mol L ⁻¹ with the tropics equation: Δ_iP = 3.40C + 3.43 × 10⁻⁶, and the detection limit is 1 × 10⁻⁸ mol L⁻¹. And the iron-porphyrin-SWNT-DMF modified GCE was applied successfully in the determination of Sudan I in real samples.     

Pico molar assay of riboflavin in human urine using voltammetry

Using a new type of DNA and carbon nano tube (CNT) mixed paste electrode using cyclic and square wave anodic stripping voltammetric (SWASV) methods, this study presents an assay of riboflavin (RF) under optimum conditions. Results of the experiment yielded a low working concentration range of nanograms with 1–10 and 10–170 ng L⁻¹ and 5–105 μg L⁻¹, at an accumulation time of 80 s in a 0.1 M H₃PO₄ electrolyte solution. A relative standard deviation of 30 μg L⁻¹ was observed at an accuracy level of 0.1164% (n = 15) under optimum conditions. The detection limit (S/N) was pegged at 0.2 ng L⁻¹ (5.31 × 10⁻¹³ mol L⁻¹ RF). The proposed method was successfully applied to an actual human urine and drug sample, and can be applied to assays of other biological samples.     

Determination of polycyclic aromatic hydrocarbons in edible oils and barbecued food by HPLC/UV–Vis detection

Determination of nine polycyclic aromatic hydrocarbons in corn, sunflower, olive oils and barbecued meat and fish by HPLC/UV–Vis method is described. The extraction procedure included a saponification, liquid–liquid extraction and finally purification of PAHs through a house-made silica–alumina column. Chromatographic determination was based on separation of PAHs on ODS column and measurement at 254 nm. All polycyclic aromatic hydrocarbons were separated and analyzed in 12 min on reversed phase ODS column with acetonitrile/water mobile phase at 1.5 mL min⁻¹ flow rate. The detection limits of nine polycyclic aromatic hydrocarbons ranged from 0.26 to 1.15 μg L⁻¹ at a signal/noise ratio of 3. The linearity of the method was between 0.9951 and 0.9996. Oil samples contain different PAHs ranging from 0.44 to 98.92 μg L⁻¹. Barbecuing process increased the concentration (in the range of 2- to 8-fold) and caused the formation of PAHs in food samples.     

Trace element levels in honeys from different regions of Turkey

A survey of 25 honey samples from different botanical origin, collected all over the Turkey was conducted to assess their trace element contents. The aim of this study was to determine the levels of cadmium (Cd), lead (Pb), iron (Fe), manganese (Mn), copper (Cu), nickel (Ni), chromium (Cr), zinc (Zn), aluminium (Al) and selenium (Se) in honey samples from different regions of Turkey. Trace element contents were determined by a flame and graphite furnace atomic absorption spectrometry technique after dry-ashing, microwave digestion and wet-digestion. The accuracy of the method was corrected by the standard reference material, NIST-SRM 1515 Apple leaves. The contents of trace elements in honey samples were in the range of 0.23–2.41 μg g⁻¹, 0.32–4.56 μg g⁻¹, 1.1–12.7 μg g⁻¹, 1.8–10.2 μg g⁻¹, 8.4–105.8 μg kg⁻¹, 2.6–29.9 μg kg⁻¹, 2.4–37.9 μg kg⁻¹, 0.9–17.9 μg kg⁻¹, 83–325 μg kg⁻¹ and 38–113 μg kg⁻¹ for Cu, Mn, Zn, Fe, Pb, Ni, Cr, Cd, Al and Se, respectively. Iron was the most abundant element while cadmium was the lowest element in the Turkish honeys surveyed. The results showed that trace element concentrations in the honeys from different regions were generally correlated with the degree of trace element contamination of the environment.     

Occurrence of patulin and its dietary intake through apple juice consumption by the Spanish population

A survey was conducted to determine levels and dietary intake of Patulin (PAT) from apple juices consumed in Spain. One hundred samples of apple juice were bought from distinct supermarkets. PAT was extracted by a liquid–liquid extraction technique and analysed with a micellar electrokinetic chromatography (MEKC) method. 66% of the samples contained PAT over the limit of detection of the method (0.7 μg L⁻¹). The PAT apple juice mean and median levels obtained were 19.4 and 4.8 μg L⁻¹, respectively, in a range between 0.7 and 118.7 μg L⁻¹. In 11% of the samples, PAT contamination exceeded the maximum permitted level of 50 μg L⁻¹ established by the EU regulation. In Spain, no significant variations were observed with respect to data published 15 years ago.     

Solid phase extraction with flame atomic absorption spectrometry for determination of traces of Ca,K, Mg and Na in quality control of white sugar

A fast and straightforward pre-concentration procedure based on solid phase extraction with a strongly acidic cation-exchanger Dowex 50 W × 8–400 was proposed to determine traces of Ca, K, Mg and Na in white sugar samples by means of flame atomic absorption spectrometry. For this purpose, 20% (m/v) white sugar solutions (100 ml) were driven through resin beds at 10 ml min⁻¹ to retain Ca, K, Mg and Na ions and to separate sucrose that passed through unretained. Thereafter, columns were rinsed with water and elements of interest were recovered prior to measurements using 5 ml of a 2 mol l⁻¹ HCl solution. Detection limits of 0.04, 0.05, 0.02 and 0.01 μg g⁻¹ for Ca, K, Mg and Na, respectively, and precision of measurements within 1–3% were achieved. The proposed method enabled to determine Ca, K, Mg and Na in samples of white sugar within corresponding ranges: 0.66–0.99 μg g⁻¹ (Ca), 2.9–12.2 μg g⁻¹ (K), 0.53–1.57 μg g⁻¹ (Mg) and 0.06–0.30 μg g⁻¹ (Na). Accuracy of this sample pre-treatment procedure and analysis method was assessed by performing spikes and recovery experiments. Recoveries of added Ca, K, Mg and Na were found to be within 97–102%, demonstrating good reliability of results.     

Total content of As,Sb, Se,Te and Bi in Spanish vegetables,cereals and pulses and estimation of the contribution of these foods to the Mediterranean daily intake of trace elements

As, Sb, Se, Te and Bi were quantified in vegetables, pulses and cereals, in order to increase the available information on the presence of these elements in the Mediterranean daily intake. Samples were dry ashed and the ashes dissolved with diluted HCl. Hydride generation atomic fluorescence spectrometry was used for quantification. Limits of detection (LOD) ranged from 0.2 to 1.7 ng g⁻¹ and the accuracy was assessed by the analysis of three certified reference materials, Rice Flour (NIST 1568a), Tomato Leaves (NIST 1573) and Cabbage (IAEA 359). Good concordance between results found and certified values were always observed. As, Sb, Se, Te and Bi levels varied in samples analysed from values under the LOD till 1001, 47, 270, 104 and 48 ng g⁻¹, respectively.     

Effect of modifiers for As,Cu and Pb determinations in sugar-cane spirits by GF AAS

Palladium plus magnesium nitrates with and without Ir, Ru and W were evaluated for the simultaneous determination of As, Cu and Pb in cachaça by graphite furnace atomic absorption spectrometry. For 20 μL of sample, 5 μL Pd(NO₃)₂ and 3 μL Mg(NO₃)₂ dispensed together onto the Ir-coated platform of the THGA, analytical curves in the 0–30.0 μg L⁻¹ As, 0–1.50 mg L⁻¹ Cu and 0–60.0 μg L⁻¹ Pb were built up and typical linear correlation coefficients were always better than 0.999. The limit of detection was 1.30 μg L⁻¹ As, 140 μg L⁻¹ Cu and 0.90 μg L⁻¹ Pb. As, Cu and Pb contents in 10 cachaça samples agreed with those obtained by ICP-MS. Recoveries of spiked samples varied from 96% to 106% (As), 97% to 112% (Cu) and 92% to 108% (Pb). The relative standard deviation (n = 12) was typically 2.7%, 3.3% and 1.9%.     

Nitrate-N determination in leafy vegetables: Study of the effects of cooking and freezing

Nitrate upon reduction to nitrite can cause methaemoglobinaemia or act as precursor in the endogenous formation of carcinogenic nitrosamines. The leafy vegetables are the major vehicle for the entry of nitrate into the human system. The present study was conducted to establish a flow injection analysis (FIA) technique to investigate the nitrate-N contents of four commonly consumed fresh leafy vegetables (Chinese cabbage, celery, lettuce and English cabbage) from market in Fiji. Two extraction techniques (activated carbon and alkaline extraction) were assessed to extract nitrate-N and the activated carbon extraction was preferred over alkaline extraction and applied. The recoveries of spiked nitrate-N in vegetable matrices ranged from 90.40% to 112.80% in activated carbon extraction with an average of 100.62%. The effects of cooking (boiling, baking and frying) and deep-freezing on the nitrate-N contents were also studied. Nitrate contents in selected leafy vegetables were determined by FIA coupled with Greiss protocol involving sulfanilamide and N-(1-naphthyl)ethylenediamine dihydrochloride as color reagents. Nitrate was determined in the linear range from 1.0 to 20.0 mg L⁻¹ with the method detection limit of 0.042 mg L⁻¹ (0.34 mg kg⁻¹). The results of the study show that nitrate contents in fresh leafy vegetables ranged from 1297 to 5658 mg kg⁻¹. Boiling reduces nitrate content by 47–56% whereas frying in Soya bean oil elevates nitrate content by as much as 159–307%. No significant change was observed in nitrate content after baking. The deep-freezing of the selected leafy vegetables shows that nitrate-N content fluctuates slightly from the original nitrate-N values over the seven day period. The FIA throughput was 38 samples h⁻¹.     

Method development for sensitive determination of nisin in food products by micellar electrokinetic chromatography

A rapid and sensitive micellar electrokinetic chromatography (MEKC) method has been developed for the identification and quantification of nisin in food products. Factors such as micelle concentration, pH, and concentration of the buffer were investigated in order to determine the optimum conditions for the analysis. Using optimised conditions of a background electrolyte containing 50 mM sodium phosphate, 80 mM sodium dodecyl sulphate (SDS) at pH 3.75 enabled the successful detection of nisin within 6 min. Limits of detection (LOD) and quantification (LOQ) were in the range of 0.3–0.8 and 1.0–2.8 mg L⁻¹, respectively. The calibration curves were linear for nisin concentration over the range of 2–60 mg L⁻¹. The relative standard deviation (RSD) of the peak area ratios and migration times for method repeatability (n = 3) were found to be lower than 5% and 1%, respectively. The potential of the proposed method to be used for quantitative determination of nisin concentrations in food materials was demonstrated by spiking food samples including dairy products, salad dressings, alcoholic beverages and canned tomatoes with known concentrations of nisin. Quantitative recoveries ranging from 83% to 104% were obtained in the food matrices.     

Optimization of ethanol extraction and further purification of isoflavones from soybean sprout cotyledon

Isoflavones from cotyledons of soybean sprouts were extracted with aqueous ethanol and further concentrated to obtain a product with a high concentration of isoflavone. The ethanol concentration, extraction time and reaction temperature were optimized by using response surface methodology (RSM). Isoflavones in aqueous ethanol were concentrated by a three-step procedure comprised of solid phase extraction (SPE) with Diaion HP-20 and Amberlite-XAD-2 adsorption columns, acid hydrolysis, and liquid–liquid extraction. The maximum amount of isoflavone in aqueous ethanol extracts (11.6 mg/g solid) was obtained when isoflavones in cotyledons (2.18 mg/ g solid) were extracted with 80–90% (v/v) aqueous ethanol above 90 °C for more than 100 min. The isoflavone extracts, obtained by SPE with a Diaion HP-20 column contained 100 mg/g solid. The liquid–liquid extraction (LLE) with ethyl ether further concentrated the extracts up to 229 mg/g solid, retaining 63% of the initial isoflavones.     

Feasibility and application of liquid–liquid extraction combined with gas chromatography–mass spectrometry for the analysis of phenolic acids from grape polyphenols degraded by human faecal microbiota

In this study the feasibility of a LLE–GC–EI-MS method for the analysis of 43 phenolic acids belonging to different chemical structure families which have been described in the literature as microbial-derived metabolites after consumption of dietary polyphenols was proved. In addition, the method was applied for the characterisation of phenolic metabolites resulting from the incubation, in anaerobic conditions, of a commercial grape seed extract (GSE) and their corresponding flavan-3-ol monomeric (GSE-M) and oligomeric (GSE-O) fractions with human faeces from healthy volunteers (n = 3). The method showed average values of repeatability and reproducibility of 5.0% and 6.3%, respectively, adequate and low detection (1.8–30.8 μg L⁻¹) and quantification limits (6.0–102.8 μg L⁻¹) and good recovery values (95%, as average value). A total of 27 phenolic acids were identified in the faecal solutions after incubation with the grape seed extracts. In general, faecal samples incubated with GSE and GSE-M (monomeric fraction) yield a higher formation of phenolic acids compared to the samples incubated with the oligomer fraction (GSE-O).